Isaac Harary

In vitro studies on single beating rat heart cells. I. Growth and organization.

Summary 1. Single beating cells may be prepared from the young rat heart. 2. These cells may beat for up to 40 days. 3. The process of dedifferentiation is seemingly accompanied by loss of striations and cessation of beating. 4. The histological appearance was discussed. 5. Single independently beating cells grow into synchronously beating nets of cells and eventually develop into beating fibers. Synchrony depends upon protoplasmic contact. 6. Two types of cells are seen, independent “leading” cells which may beat spontaneously and “following” cells which beat only upon contact with beating cells.

In vitro Studies of Single Isolated Beating Heart Cells

Rat heart cells, separated by trypsin treatment and grown attached to glass in a liquid medium, exhibit periodic contractions similar to a whole beating heart. The rate of beating, which is up to 150 beats per minute, is affected by cardiac drugs and by metabolic substrates and inhibitors.

In Vitro studies on single beating rat heart cells: VI. Electron micorscopic studies of single cells1

The ultrastructure of single beating heart cells, isolated from newborn rats by trypsinization and grown in semisynthetic media using a modified feeder layer technique, was investigated. In many of the cells myofibrils with A, I, and Z bands were seen. In all beating cells more or less well developed myofilaments were seen. Varying developmental stages occurred in cultures of the same age. Well developed mitochondria and Golgi bodies were also present. Ribosomes were found linearly arranged parallel to the myofilaments organized in what appeared to be a helix. Polysomes, scattered filaments, and Z material seem to arise in the periphery of the cell. The relation of these polysomes to the synthesis of myofilaments and the development of myofibrils is discussed.

In vitro studies on single beating rat heart cells. II. Intercellular communication.

Summary 1. Rat heart cells separated by trypsin and grown in vitro continue to beat. 2. Cells beating at an independent rate become synchronous upon physical contact, indicating that the communication of rate of beating is through protoplasmic contact. 3. The rate of beating for the synchronous net of muscle cells is determined by the fastest beating cell. 4. Despite the superimposed rate, each cell retains potentially its own inherent rate of beating.

Studies in vitro on single beating heart cells VIII. The effect of oligomycin, dinitrophenol, and ouabain on the beating rate

Summary 1. Dinitrophenol inhibits beating in cultured heart cells and ATP restores the beating. 2. A combination of iodoacetate and oligomycin is necessary to inhibit beating, while either alone is insufficient. 3. Oligomycin can restore beating of dinitrophenol-inhibited cells. 4. These results indicate that ATP from either glycolysis or oxidative phosphorylation may support beating in cultured heart cells. 5. High-energy intermediates synthesized before the site of oligomycin inhibition are not able to maintain beating.

Studies on the effects of essential fatty acids on growth rate, fatty acid composition, oxidative phosphorylation and respiratory control of HeLa cells in culture

Abstract 1. Mammalian cells in culture provide a simple model to study the function of polyunsaturated fatty acids. 2. HeLa S 3 cells cultured in a lipid-deficient medium showed a pattern of essential fatty acid deficiency represented by an increase in 18:1 and a decrease in 20:4 and 18:2, a lack of growth and impaired mitochondrial function. 3. Addition of albumin-bound linoleic or arachidonic acid partially or totally prevented these changes.

[74] The isolation and cultivation of rat heart cells☆

Publisher Summary Heart cells are prepared by dissociation of intact 2–5-day-old rat hearts and cultivation of the component cells attached to a sterile surface in a suitable medium. After 1 or 2 days, the successful culture contains a majority of cells that demonstrate their myocardial nature by contracting spontaneously. As in the preparation of all primary cultures, one must be prepared to face variations, in the resultant cell population that are not easy to control. Although the majority of cells in the culture are muscle cells, there exist variable amounts of endothelial and fibroblast cells. In this case the percentage of functioning muscle cells can be determined visually, by the number of beating cells; histochemically, by the number of cells containing myosin or other muscle proteins; or biochemically, by measuring the total myosin content in the culture dish. The variability can arise from the differences in the serum, and dissociating agent; and variation in the quality of the air and the state of the animals.

In vitro studies of beating-heart cells in culture. XI. The ATP level and contractions of the heart cells

Abstract 1. The beating of heart cells in tissue culture is not affected by the inhibition of either glycolysis or oxidative phosphorylation alone. When both pathways are inhibited simultaneously, beating stops. 2. The ATP concentration does not change significantly when either glycolysis or oxidative phosphorylation is inhibited alone. When both glycolytic and oxidative phosphorylation inhibitors are present, the ATP level drops significantly. The combined effect of both inhibitors exceeds the sum of the effects of the individual inhibitors. 3. Dinitrophenol inhibits the beating of heart cells with a concomitant small decrease in ATP level. The addition of oligomycin restores the ATP level to control values and beating resumes. 4. Heart cells in which beating is inhibited, respond to electrical stimulation, even though the ability to beat spontaneously has been lost. Contractions in response to stimulation can be observed until the decrease of ATP has reached 85–90%. 5. It is concluded that heart cells must have a mechanism for maintaining a high steady-state level of ATP, since such a high level is required for spontaneous beating. The possible mechanisms for the maintenance of high steady-state level of ATP are discussed.

IN VITRO STUDIES ON SINGLE BEATING RAT HEART CELLS. VII. ULTRASTRUCTURE OF THE BEATING CELL LAYER.

The ultrastructure of synchronously beating heart cells grown in culture was observed. The cells appeared to vary as to the stage of development as judged by the appearance of the myofilaments and mitochondria. In the area of contact between cells, cell membrane thickenings, attached to the myofibrils, were seen. The relation of these structures to the development of intercalated disks and myofibrils and the propagation of excitation in the synchronous cells was discussed. Cells whose centers are crowded with radially arranged fibrils were observed, and their possible relation to their function as pacemaker or “leading” cells was discussed. The presence of “knot”-like structures with Z bands and fibrils was related to their possible function as centers of contraction in the cell.

Studies in vitro on single beating rat-heart cells X. The effect of linoleic and palmitic acids on beating and mitochondrial phosphorylation

Abstract 1. Beating rat-heart cells cultured in a lipid-deficient medium showed: (a) lack of growth; (b) decrease and later a cessation of the beating; (c) impairment of oxidative phosphorylation and a decrease in the respiratory control; (d) a pattern of fatty acid deficiency as shown by the analysis using gas-liquid chromatography. 2. The addition of albumin-bound linoleic acid or arachidonic acids prevented partially or totally the effects of the lipid deficiency on the mitochondrial function and the fatty acid composition, but did not promote growth and did not improve the beating. 3. Medium supplemented with albumin-bound palmitic acid maintained the beating, but did not promote growth and did not affect the impairment of the mitochondrial function.