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J W Greenawalt

Publications and source records attributed to J W Greenawalt.

At least 19 recordsLinked to original sources
PubMed1978

Transport of 5-hydroxytryptamine by dense granules from porcine platelets.

A method is described for the isolation of a homogeneous preparation of dense granules from procine platelets. The purified dense granule fraction contained approximately 400 nmol of 5-hydroxytryptamine/mg of protein and appeared to be homogeneous when examined by electron microscopy. Isolated dense granules transport exogenously added 5-hydroxytryptamine via two mechanisms: 1) a carrier-mediated process predominating at low substrate concentrations and 2) a diffusion-controlled process predominating at high substrate concentrations. Temperature studies revealed an apparent energy of activation of 14.9 kcal/mol for the carrier-mediated transport. Kinetic data yielded a Km of 3.3 micron and a Vmax of 0.79 nmol/min/mg of protein for the mediated transport process. Steady state uptake was sensitive to changes in medium osmotic pressure and a decline in uptake below 300 mosM was correlated with release of endogenous 5-hydroxytryptamine. The transport was inhibited by a number of structural analogs of 5-hydroxytryptamine. These results demonstrate the existence of a carrier-mediated transport system for 5-hydroxytryptamine in the membranes of the platelet dense granules.

Animals↗
PubMed1977

Composition and synthesis of cellular lipids in Neurospora crassa during cellular differentiation.

The synthesis of cellular lipids of Neurospora crassa was measured during growth on low (2% sucrose)- and high (15% glucose)-carbohydrate supplementation. The amount of lipid per dry weight of cells does not change during the germination and early logarithmic growth periods, but the percentage of phospholipid in the lipid does increase, reaching a maximal value of 90% at 4 to 5 h after inoculation, at which time the phospholipid content of the cells is approximately 60 mumol/g (dry weight). The content of the anionic phospholipids, as a percentage of the lipid fraction, is relatively constant during the growth period, but the contents of the zwitterionic phospholipids phosphatidylcholine and phosphatidylethanolamine change in a reciprocal fashion. During the first 8 h of growth, phosphatidylcholine falls from 53% of the phospholipid to 43%, whereas phosphatidylethanolamine rises from 29 to 38%. The total of these two phospholipids is approximately 83% during the growth period studied. The synthesis of cellular phospholipids, measured either by [32P]H3PO4 or [14C]glucose incorporation, reached maximal levels between 3 and 5 h of growth. The effect of the high-carbohydrate supplement on cellular lipids was minimal. Inclusion of 15% glucose decreased the labeling of phospholipid by [32P]H3PO4, but did not affect lipid composition. This observation is in contrast to the effects of high glucose on mitochondrial phospholipid synthesis.

Cell Differentiation↗
PubMed1976

Biogenesis of mitochondrial membranes in Neurospora crassa during cellular differentiation: changes in oxidative phosphorylation and synthesis of mitochondrial phospholipids.

Changes in the capacity of mitochondria to carry out oxidative phosphorylation and in the rate of synthesis and incorporation of phospholipids into mitochondria were measured during the germination of conidiospores of Neurospora crassa. The competence of isolated mitochondria to carry out coupled respiration was very low during the first 3 h growth, but it increased rapidly, reaching maximal levels at 5 to 6 h growth. Changes in mitochondrial function were the same in cells grown in 2% sucrose- or 15% glucose-supplemented medium. The rate of synthesis of mitochondrial phospholipids was very low during the first 2 h growth and increased to maximal levels between 3 and 5 h. The rate of synthesis of mitochondrial phospholipids was approximately three times higher in cells grown in 15% glucose than in those grown in 2% sucrose. The maximal rate of synthesis of mitochondrial phospholipids occurred during spore germination and preceded attainment of full competence for oxidative phosphorylation. The lipid-rich condition of the mitochondrial resulting from the high rate of synthesis of phospholipids in glucose-grown cells is postulated to be related to the whorled inclusions observed in thin sections of Neurospora cells.

Antimycin A↗
PubMed1976

Biogenesis of mitochondrial membranes in Neurospora crassa during cellular differentiation: ultrastructural changes accompanying differentiation.

The ultrastructural characteristics of Neurospora cells during dedifferentiation and redifferentiation of conidiospores into vegetative cells have been determined. This germination process occurs between 2 and 5 h after inoculation; by 3-5 h, approximately 50% of the cells have germinated. The cells enter the exponential phase of dry-weight gain between 4 and 5 h after inoculation. Several unusual structures are observed in Neurospora cells during germination. Whorled structures are frequently seen in the cytoplasm during germination, and occasionally at other times. They appear to be derived from the cytoplasmic membrane. Whorled structures of different appearance were observed in the mitochondria between 2 and 4 h after inoculation. Their number was related to the level of metabolizable carbohydrate, and was higher in 15% glucose-than in 2% sucrose-supplemented medium, and very low in medium containing 15% mannitol, or 2% sucrose+13% 2-deoxyglucose, or no added carbohydrate. The mitochondrial inclusions were osmiophilic and could be removed by treatment with 90% aqueous acetone in the cold, indicating that they were composed at least in part of lipid. The strong dependence of the number of mitochondrial inclusions on time and on carbohydrate supplementation, suggests that there is a physiological basis for these structures and that they reflect changes occurring in the mitochondria at times significant to cellular differentiation.

Carbohydrate Metabolism↗
PubMed1975

Biogenesis of mitochondrial membranes in Neurospora crassa. Mitochondrial protein synthesis during conidial germination.

The conidia of Neurospora crassa entered logarithmic growth after a 1-h lag period at 30 degrees C. Although [14C]leucine is incorporated quickly early in growth, cellular protein data indicated that no net protein synthesis occurred until after 2 h of growth. Neurospora is known to produce ethanol during germination even though respiratory enzymes are present. Also, Neurospora mitochondria isolated from cells less than 3-h old are uncoupled. Since oxygen uptake increased during germination, was largely cyanide-sensitive, and reached a maximum at 3 h, it is hypothesized that during early germination the uncoupled electron transport chain merely functions to dispose of reducing equivalents generated by substrate level ATP production. The rate of protein synthesis in vitro by mitochondria isolated from 0-8-h-old cells increased as did cell age. Mitochondrial protein synthesis in vivo, assayed in the presence of 100 mug cycloheximide/ml, increased from low levels in the cinidia to peak levels at 3-4 h of age and then slowly decreased. The rate of mitochondrial protein synthesis in vivo was linear for at least 90 min in 0-4-h-old cells, but declined after 15 min of incorporation in 6 and 8-h-old cells. The products of mitochondrial protein synthesis in vivo were analyzed with dodecylsulfate gel electrophoresis and autoradiography. Early in germination 80% of the synthesis was of two small proteins (molecular weights 7200 and 9000). At 8 h 85% of the radioactivity was in 10 larger proteins (12 200 to 80 000). Within the high-molecular-weight class, proteins of between 12 000 and 21 500 molecular weight were preferentially lavelled early in germination, whereas after 8 h of growth proteins of 27 500 to 80 000 molecular weight were preferentially labelled. It is hypothesized that the 7200 and 9000-molecular-weight products of mitochondrial protein synthesis combine with other proteins to form the larger proteins found later in growth. The availability of these other proteins in cells of different ages could affect the rate of mitochondrial protein synthesis in vivo.

Adenosine Triphosphate↗
PubMed1974

Biochemical and ultrastructural aspects of Ca2+ transport by mitochondria of the hepatopancreas of the blue crab Callinectes sapidus.

Mitochondria isolated from the hepatopancreas of the blue crab Callinectes sapidus show up to 12-fold stimulation of respiration on addition of Ca(2+), which is accompanied by Ca(2+) accumulation (Ca(2+):site = 1.9) and H(+) ejection (H(+):Ca(2+) = 0.85). Sr(2+) and Mn(2+) are also accumulated; Mg(2+) is not. A strongly hypertonic medium (383 mosM), Mg(2+), and phosphate are required for maximal Ca(2+) uptake. Ca(2+) uptake takes precedence over oxidative phosphorylation of ADP for respiratory energy. Once Ca(2+) is accumulated by the crab mitochondria, it is stable and only very slowly released, even by uncoupling agents. ATP hydrolysis also supports Ca(2+) uptake. Respiration-inhibited crab hepatopancreas mitochondria show both high-affinity and low-affinity Ca(2+)-binding sites, which are inactive in the presence of uncoupling agents. Crab hepatopancreas mitochondria have an enormous capacity for accumulation of Ca(2+), up to 5,500 ng-atoms Ca(2+) per mg protein, with an equivalent amount of phosphate. Freshly isolated mitochondria contain very large amounts of Ca(2+), Mg(2+), phosphate, K(+), and Na(+); their high Ca(2+) content is a reflection of the vary large amount of extra-mitochondrial Ca(2+) in the whole tissue. Electron microscopy of crab mitochondria loaded with Ca(2+) and phosphate showed large electron-dense deposits, presumably of precipitated calcium phosphate. They consisted of bundles of needle-like crystals, whereas Ca(2+)-loaded rat liver mitochondria show only amorphous deposits of calcium phosphate under similar conditions. The very pronounced capacity of crab hepatopancreas mitochondria for transport of Ca(2+) appears to be adapted to a role in the storage and release of Ca(2+) during the molting cycle of this crustacean.

Adenosine Diphosphate↗