Hypertonic sucrose

Philos Trans R Soc Lond B Biol Sd 354 337-46 Khvotchev M, Lonart G, Sudhof TC (2000) Role of caldum in neurotransmitter release evoked by alpha-latrotoxin or hypertonic sucrose. Neuroscience 101 793-802 Klenchin VA, Kowalchyk JA, Martin TF (1998) Large dense-core vesicle exocytosis in PC12 cells. Methods 16 204-8... 

The ability of a-LTX to trigger neurotransmitter exocytosis in the absence of extracellular Ca2+ remains particularly interesting and inexplicable to the field (Longenecker et al. 1970 Ceccarelli et al. 1979 see also Siidhof (2001) and Ushkaryov et al. (2004) for review). This is clearly different from depolarization-induced exocytosis, which is Ca2+-dependent, but not unlike the effect of hypertonic sucrose. The possibility that a-LTX-induced release involves an unknown, Ca2+-independent mechanism which may also occur during normal synaptic activity has provided the casus belli for many a quest for a-LTX structure and receptors that could trigger neurotransmission via intracellular mechanisms. 

The characteristics of Ca2+-independent release are peculiar it requires the presence of divalent cations, such as Mg2+, which can be added or removed in succession, causing respective bouts of secretion or its cessation (Misler and Hurlbut 1979). In the absence of Mg2+, this release can be supported by slightly hypertonic sucrose, by itself insufficient to cause secretion (Misler and Hurlbut 1979). The Ca2+-independent release can be blocked by millimolar La3+ (Rosenthal et al. 1990) or concanavalin A (Grasso et al. 1978 Boehm and Huck 1998). It may involve release of Ca2+ from mitochondria, as observed in peripheral (Tsang et al. 2000) and not central synapses (Adam-Vizi et al. 1993), but it is unclear if stored Ca2+i itself can trigger release. 

Slavicek J. 1972. Effect of Ba2+ on contractility of the isolated right rat ventricle. Substitution of NaCI for chlorine of hypertonic sucrose. Physiol Bohemoslov 21 189-199. 

Fluorescent cholera toxin Fluorescent dextran conjugates Fluorescent Dil-LDL Fluorescent fusion proteins (GFP) Fluorescent / neutralizing IgG Fluorescent transferrin Hypertonic sucrose Ikarugamycin... 

Clathrin-mediated endocytosis can be blocked by several pharmacologic inhibitors, including the antipsychotic drug chlorpromazine (Thorazine), the natural product ikarugamycin, and the antiviral drug amantadine. The metabolic poisons phenylarsine oxide and sodium azide also block CMF but additionally inhibit protein synthesis. Culture of cells under conditions that deplete potassium or calcium, treatment of cells with hypertonic sucrose, or acidification of the cytoplasm by addition of... 

Ox liver mitochondria were swollen in hypotonic Tris buffer and then were shrunk in hypertonic sucrose. This technique liberated a mitochondrial glycoprotein which was purified by preparative electrophoresis... 

This conclusion is based on immunogold localization of PrP in these organelles by electron microscopy inhibition of PrP internalization by incubation of cells in hypertonic sucrose, which disrupts clathrin lattices and detection of PrP in purified preparations of coated vesicles from brain. The N-terminal half of the PrP polypeptide chain is essential for efficient clathrin-mediated endocytosis, because... 

Skeletal Muscle DNA. Hypertonic sucrose, bupivicaine, Naja nagricollis cardiotoxin, gene gun, jet injector... 

The periplasmic contents are released by a mild cold osmotic shock procedure. In essence this consists of plasmolysing the bacteria by hypertonic sucrose/EDTA, followed by resuspension in cold distilled water. This releases approximately 4 per cent of the total cell protein the bacteria remain viable, though unable to perform any function dependent upon the lost periplasmic binding protein. 

Evidence to substantiate this concept was immediately forthcoming. Investigations by Claude and by Schneider and their collaborators indicated that the succinoxidase and cytochrome oxidase activities of liver cells are largely concentrated in the mitochondria, a finding which explained the results of earlier studies on these insoluble enzymes. These workers also developed a technique for the fractionation of tissue extracts in hypertonic sucrose solution, a technical advance of considerable value, since previous work with distilled water or buffered saline extracts did not preserve the morphological and enzymological properties of the fractions satisfactorily. With this method, it was possible to isolate mitochondria from liver tissue in a state approximating that found in the intact cell, with the full preservation of many labile enzyme activities which it had been impossible to study by previous methods. 

Working with hypertonic sucrose homogenates, Kennedy and Lehninger found that isolated washed mitochondria of rat liver tissue oxidized not only succinate but also other key intermediates of the Krebs cycle. They also reported that fatty acid oxidation was localized in the mitochondria and furthermore demonstrated that phosphorylations coupled to these oxidations took place in these preparations, as could be shown with tracer phosphate experiments. 

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