Mouse from dissociated

The implementation of animal test protocols in the 1980s has been accompanied by the development of a host of alternative methods to study adverse effects of chemicals on reproductive and developmental parameters. For example, rat whole embryo culture stems from the seventies (16), as does the rat limb bud organ culture (17) and rat limb bud and brain micromass was developed in the eighties (18). An elegant nonvertebrate alternative model used regeneration of polyps of Hydra atUnuata from dissociated cells (19). Animal-free in vitro alternatives include those employing the proliferation of a human embryonic palatal mesenchymal cell line (20), the attachment of a mouse ovarian tumor cell line (21), and the differentiation of a neuroblastoma cell line (22) and a embryonal carcinoma cell line (23). Various overviews of methods have been published over the years (24). The predictability of... 

Schousboe A, Svenneby G, and Hertz L (1977) Uptake and metabolism of glutamate in astrocytes cultured from dissociated mouse brain hemispheres / Neurochem 29, 299. 

For extracellular signal recordings from electrically active cells in culture, two main concepts have been developed in the past microelectrode arrays (MEAs) (see Fig. 2a) with metalized contacts on silicon or glass substrates have been used to monitor cardiac impulse propagation from dissociated embryonic myocytes [1-3], dissociated invertebrate neurons [4, 5] and mammalian neurons [6] spinal cord [7], and mouse dorsal root... 

Leptin is a cytokine produced and secreted by adipose tissue in proportion to the body fat content [3]. Mice and humans lacking leptin or its receptor develop a severe hyperphagia and a dramatic degree of obesity which is considerably more pronounced than that of the NDRKO mouse. Thus, leptin is the key adiposity signal in rodents and humans. Leptin secretion appears to reflect the metabolic status of the adipocyte rather than the sheer size of triglyceride deposits, and leptin levels may transiently be dissociated from total body fat. Nonetheless, over the course of a day with unrestricted food supply, plasma leptin levels reliably reflect the amount of total body fat. Local administration of leptin into the brain results in reduced food intake. The vast majority of patients with obesity have elevated serum levels of leptin. Thus, it is believed that the polygenic obesity is due to leptin resistance rather than to inadequate leptin secretion, or to a reduced blood/brain transport of the cytokine. 

Homogenates of MetruUum senile, possibly the world s most common large sea anemone, yield extracts that are powerfully hemolytic for washed mammalian erythrocytes (22). The active substance, metridiolysin, is a protein of molecular weight approximately 80,000. In contrast to the sphingomyelin-inhibitable toxins, metridiolysin is an acidic protein having a pi of about 5. It is thermolabile and is inactivat by proteolytic enzymes. The optimal pH for hemolysis is between 5 and 6, and at pH 8 the lysin is inactive. It can be dissociated into two subunits of unequal size. Besides being cytolytic in vitro, metridiolysin is lethal when injected intravenously into mice. As shown in Table IV erythrocytes from the horse or dog are about a hundred times as sensitive to lysis as those from the mouse, and erythrocytes from other animals tested are intermediate in sensitivity. 

Froscio SM, Humpage AR, Burcham PC, Falconer IR (2003) Cylindrospermopsin-induced protein synthesis inhibition and its dissociation from acute toxicity in mouse hepatocytes. Environ Toxicol 18 243-251... 

Gould TW, Buss RR, Vinsant S, Prevette D, Sun W, Knudson CM, MilUcan CE, Oppenheim RW (2006) Complete dissociation of motor neuron death from motor dysfunction by Bax deletion in a mouse model of ALB. J Neurosc. 26(34) 8774-8786... 

Wang Y, Osterbur DL, Megaw PL et al 2001 Rhythmic expression of Nocturnin mRNA in multiple tissues of the mouse. BMC Dev Biol 1 9 Welsh DK, Logothetis DE, Melster M, Reppert SM 1995 Individual neurons dissociated from rat suprachiasmatic nucleus express independently phased circadian firing rhythms. Neuron... 

Most of the data is taken from earlier experimental work. The concentrations in the feed and reference values are taken from mouse and rat brain data for the sake of illustration. The membrane permeability parameters for S2 and S3 are assumed equal to the value for si. The normalized parameter > is taken equal to I>2, which was found earlier experimentally. The kinetic parameters 6m for m = 1,.., 8 of reaction (1) are chosen by using a known dissociation constant and by keeping the experimentally found proportion for reaction (2) on substrate-inhibition and hydrogen-ion effects. 

Our initial studies were carried out on cultures of dissociated cells from brains of one day old mice. Unfortunately these preparations had lower than hoped for activities of certain biochemical parameters of myelination. For example, both the incorporation of 35g0jj into sulfolipids and the activity of the cyclic nucleotide phosphohydrolase attained relatively small activities even at the age (8 to 11 days) of optimum activity in culture (fig. 1 and 2). The dissociated cells from the 1 day old mouse produced parallel developmental patterns of both myelin parameters with peak activities at 8 to 11 days in culture. However cultures prepared from 15 day fetal brains proved to have relatively... 

Figure 1. Incorporation of 3sSOi2 into lipids of dissociated cells from 15-day embryonic and from 1-day postnatal mouse brain grown for different days in vitro. The 35SO,2 was exposed to the cells for 16 hr (a time in which the incorporation of 3SS still proceeded linearly). The data are expressed as activity from 35SO,2 incorporated into the total number of cells (or total proteins) per flask (21).

Figure 3. Time course of incorporation of ssSOi>" into cerebroside sulfate (cer-SOk) and sulfogalactosyl glycerol lipid (SGG-lipid) in dissociated brain cells from 15-day mouse embryos grown 19 days in culture...

Table III gives the results of an experiment in which the effect of serum manipulation on sulfolipid synthesis by dissociated brain cells was examined. The cells isolated from the embryonic mouse brain were grown on the medium containing calf serum (20%) for 3 days by which time most of the cells would have attached to the substratum. On the 4th day, the medium was replaced by fresh medium containing calf serum, calf serum + 13(2 x 10 8m), hypothyroid calf serum, or hypothyroid calf serum + 13(13 ng/ml). Cultures were grown for another week and then labeled with 400 pCi for 16 h. The lipids...

Figures for K, the dissociation constant of the enzyme-inhibitor complex, obtained under strictly comparable conditions with mouse-liver preparations, are shown in Table VIII for D-glucaro-1,4-lactone and some closely-related compounds, all of which act competitively. Table IX gives figures obtained with the enzyme from female-rat preputial gland and various substrates within the limits of these experiments, Kt for boiled D-glucarate or boiled galactarate solutions was little affected by changes in the purity of the enzyme or the pH of test. In accordance with theory, similar values of Ki for any one inhibitor were obtained with different substrates, with the notable exception of o-nitrophenyl /8-galacturonide.

Sulfuric acid, phosphate, and sucrose extracts of mouse and beef pancreas have also been compared chromatographically on Amberlite IRC-50 (154). For both species, the sulfuric acid extracts give two peaks which correspond to ribonucleases A and B. When prepared from supernatant free of zymogen granules and microsomes, phosphate and sucrose preparations give a third peak, (10-15 % of the total activity in the case of mouse 1 % in the case of beef) which can be converted into the previous ones by acid treatment. This treatment induces a two- to three-fold increase in activity and the dissociation of the acidic compound already referred to above. The chemical structure of mouse ribonuclease(s) is still unknown. 

Formaldehyde has been shown to significantly inhibit the viability and proliferation of mouse lymphocytes. IC50 values in a 3 h exposure ranged from 1.19x 10 to 8.20 X 10 mol 1 At 1-3 mmol 1 formaldehyde concentrations, dissociated rat thymocytes showed a dose-dependent decrease in cell viability. This may have been associated with an observed reduced cellular content of glutathione or an increased concentration of cellular calcium ion. 

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