194 MUSCULAR CONTRACTION IN TISSUE-CULTURES. 



than the explanted piece, was formed. Among the cells of this new growth were 

 many mitotic figures (figs. 7, 8, and 9). As the cells migrated out they became 

 spread out more and more closely upon the under surface of the cover slip, so that 

 the edge of the growth was composed of a single layer of large, flat cells. These 

 flat cells, although somewhat distorted in so far as the position of certain cyto- 

 plasmic structures are concerned, have an advantage over those obtained by 

 means of sections, in that all the structures of the cell are present and can be 

 observed in their relations throughout the activity of the living cell. The region 

 between these outer cells and the explanted piece may be one or several cells in 

 depth. The cells here, while largely spread out, resemble more nearly the cells 

 of the normal embryo and are very little distorted laterally. 



Spontaneously contracting cells usually were found among the less spread- 

 out cells and not among those at the edge of the growth. Cultures of the amnion 

 furnished numerous rhythmically contracting smooth-muscle cells; those of the heart 

 gave rise frequently to sheets of cells contracting in co-ordination with the beat of the 

 explanted piece, and at times to a few isolated cells beating independently; while 

 from the skeletal muscle were obtained muscle buds, muscle-fibers, and myoblasts, 

 each of which were occasionally found undergoing spontaneous contraction. 



For a better understanding of the cells in tissue-culture various preparations 

 of other living muscular tissues were made, among which may be mentioned the 

 following: (1) The entire uninjured amnion of a 3, 4 or 5 day chick embryo; (2) a 2 to 

 3 day chick embryo with beating heart; (3) preparations of teased heart muscle and 

 teased skeletal muscle-fibers of chick embryos; (4) certain microscopical marine 

 copepods whose cross-striated muscle-fibers could be studied while the animal 

 remained alive; (5) the isolated sarcostyles of the insect's wing muscle; (6) thin slices 

 of the muscle-fibers from an adult dog, cat, or turtle. In addition to the above, 

 preparations were fixed and stained in various ways in order to compare the results 

 with those of other investigators. 



TYPES OF MUSCLE. 



SMOOTH MUSCLE FROM THE AMNION. 



GROWTH FROM THE AMNION IN TISSUE-CULTURES. 



The amnion, as is well known, consists of a layer of smooth-muscle cells over- 

 lying a layer of epithelial cells, in neither of which have nerve-cells or fibers been 

 satisfactorily demonstrated (fig. 1). The outgrowth from this tissue in cultures 

 did not usually form a membrane composed of the two types of cells, but instead 

 the smooth-muscle cells (SM) and the epithelial cell (E) grew out more or less 

 independently of each other (fig. 9). No nerve-fibers were found in any cultures 

 from the amnion, regardless of the age of the chick. This behavior is quite 

 contrary to that of cultures from certain other muscles (heart, stomach, and 

 intestine) in that a large percentage of cultures of the heart and intestine contain 

 nerve-fibers. The lack of growth of nerve-fibers may be taken as an indication that 

 no ganglion cells resembling those of the heart or intestines were present in the 



