Properties of Venezuelan Equine Encephalomyelitis Virus Accompanying Attenuation In Vitro

Virus obtained during serial plaque passage of the virulent parent egg seed (PES) of the Trinidad strain of Venezuelan equine encephalomyelitis (VEE) virus produced only large plaques during either 3 serial plaque passages in chick fibroblasts or 10 plaque passages in L cells, and was lethal for mice by the intraperitoneal route. Virus showing these characteristics was designated the stable large-plaque (Ls) type. In contrast, virus obtained during serial plaque passage of the attenuated 9t strain in chick fibroblasts formed only very small plaques and was not lethal for mice by the intraperitoneal route. Virus showing these properties was designated the stable small-plaque (Ss) type. Under other passage conditions, however, large-plaque virus that yielded about 90% large and 10% small plaques was obtained; this virus was designated the unstable large or Lu type because it differed from the Ls type, which yielded only large plaques. The Lu type continued to yield the same ratio of large to small plaques for several plaque-to-plaque passages. In addition, small-plaque virus that yielded both large and small plaques and that showed a reduced capability to infect mice was also recovered. This virus was designated the unstable small or Su type because it differed from the Ss type in its higher level of virulence and in its plaque-forming properties. Thus, based upon the properties of virulence for mice and plaque size, four viral types could be discerned. The evidence suggests that serial passage in cell culture imposed environmental pressures that sequentially selected the following viral types: Ls, Lu, Su, and Ss.     

Ventricular septal defect with bidirectional shunting: Mathematical considerations

Previously proposed formulae for the quantitative estimation of bidirectional shunts across ventricular septal defects require determination of the oxygen contents of mixed venous, pulmonary artery, pulmonary venous, and aortic blood. Because these formulae do not take into account the mixing of oxygenated with unoxygenated blood within the ventricles, their use must result in underestimation of shunt flows in each direction. A mathematical model for a ventricular defect is examined, in which it is assumed that mixing of blood occurs in each of six sites in the venae cavae or right atrium, right ventricle, pulmonary artery, left atrium, left ventricle, and aorta. A total of fourteen streams of blood can flow from one to another of these mixing sites. As long as complete mixing occurs in the six specified mixing sites, any degree of mixing or non-mixing of the various streams is permitted. From the equations characterizing the model, formulae are derived in which the shunt flow in each direction is expressed in terms of the oxygen contents in the six mixing sites and the fractions of blood which enter the shunt from either side without prior mixing in a ventricular mixing site. The previously reported formulae, which apply when no ventricular mixing is allowed to occur, lead to theoretical minimum values for the shunt flows in each direction. At the opposite extreme where all the shunting blood is required to mix in a ventricle before entering the shunt, formulae for maximum possible shunt flows are also obtained. The absolute values for the left-to-right and right-to-left shunt flows, which must lie somewhere between the theoretical maximum and minimum values, cannot be computed from blood gas data alone. This work was supported in part by grant HE-07563 from the National Heart Institute of the National Institutes of Health and grants-in-aid from the American and North Carolina Heart Associations and the Life Insurance Medical Research Fund. Work completed during tenure as U.S.P.H.S. post-doctoral fellow.     

Substructure of membranes in cultivated chick embryo fibroblasts

Summary Electron microscope examination of the plasma membrane of chick embryo fibroblasts cultured in vitro revealed the presence of a single osmiophilic layer about 90 Å thick and a substructure composed of ovoid sub-units associated with an amorphous component. These ovoid sub-units measured approximately 112 Å along the major axis and 75 Å along the minor axis and were composed of a central core, approximately 30 Å by 60 Å, surrounded by a peripheral component.Examination of other membranous components of these cells revealed a similar ovoid subunit structure in a single layered membrane. Differences in thickness and in the sizes of ovoid sub-units were seen in these membranes. The ergastoplasmic membranes, the outer nuclear membranes, the outer mitochondrial and the Golgi membranes were found to be the thinnest.These varied in thickness from approximately 75 Å to 80 Å. The thickest membranes seen were the inner nuclear membranes. These were approximately 100 Å thick. The dimensions of the ovoid sub-units corresponded with differences in the thickness of the various membranes. These findings support the concept of a particulate substructure of cell membranes.This work was aided by Research Grant PH 5593 from the National Science Foundation. Some of the equipment used was purchased with funds from the National Institutes of Health Grant 2TI GM 326. I wish to thank Dr. Robert M. Dougherty from the Department of Microbiology who grew and supplied me with the chick embryo fibroblast cultures used in these studies, and Mrs. Ursula Feller fer her technical assistance.     

The Plasma Phenolsulfonphthalein Index (PSPI) of Renal Function: I. Theoretical Considerations and Investigative Studies

A simple method is described for the assessment of renal function, without collection of urine. Phenolsulfonphthalein (PSP) is injected intravenously in a dosage of 1 mg./kg. body weight, and blood samples are taken at timed intervals. Only one venepuncture is required. The index is based on the rate of decrease in concentration of PSP in the plasma 15 to 35 minutes after injection, during which time this rate is exponential. The methodology and normal range, and the influence of dose, urine flow rate, age and body surface are defined. The extrarenal component of the PSP index and the distribution of PSP in body fluids are discussed.     

REVERSIBLE AND IRREVERSIBLE CHANGES IN THE FINE STRUCTURE OF NERVOUS TISSUE DURING OXYGEN AND GLUCOSE DEPRIVATION

Rabbit retinas were fixed for electron microscopy immediately after removing the eye and after incubations in a control medium and in three different deprivation media that were identical with the control except for the omission of glucose, oxygen, or both. A systematic comparison was made of the electron microscopic appearance of the different retinas with particular attention to four regions: rod inner segments, rod synapses, bipolar cell bodies, and ganglion cell myelinated axons. Retinas fixed after 1 hour of incubation in the control medium appeared virtually identical with those fixed immediately after ocular removal. Retinas deprived of oxygen and glucose for only 3 minutes showed generalized swelling of mitochondria and alterations in the structure of the synapses with loss of synaptic vesicles. Extending the combined deprivation caused further mitochondrial swelling and synaptic changes and also led to progressive swelling of the Golgi membranes and the granular endoplasmic reticulum. All these changes were almost completely reversible for up to 20 minutes but were irreversible by 30 minutes, at which time multiple discontinuities had appeared in cell and organelle membranes. Anoxia alone produced alterations similar to those found after somewhat shorter periods of the combined deprivation, whereas glucose withdrawal produced only minor changes. These electron microscopic results correlate quite well with previously reported electrophysiological measurements.