V.-The Structure of the Coloured Blood-corpuscles of Amphiuma tridactylum, the Frog, and Man. By Dr. H. D. SCHMIDT, Pathologist of the Charity Hospital, New Orleans, La.

(Taken as read before the ROYAL MICROSCOPICAL SOCIETY, April 3, 1878.)

PLATE IV.

NOTWITHSTANDING the numerous investigations which have been made for the purpose of determining the true structure of the coloured blood-corpuscles, the views on this important subject, it appears, are still varying, leaving it an object for further research and discussion.

Formerly, these bodies were almost universally regarded as true cells, consisting of a cell-membrane, enclosing the protoplasm; and this view was entertained of the round bi-concave non-nucleated blood-disks of the blood of the Mammalia, as well as of those ellipsoidal nucleated bodies met with in the blood of other classes of vertebrated animals.

EXPLANATION OF PLATE IV.

In order to enable the reader to compare the relative sizes of the bloodcorpuscles discussed in the paper, all figures, with the exception of 45, 46, and 47, have been represented enlarged by the same magnifying power, viz. 420 diameters.

FIG. 1.-Front view of a coloured blood-corpuscle of Amphiuma tridactylum. FIG. 2.-Side view of the same.

FIGS. 3 to 12.-Various forms of coloured blood-corpuscles of the Amphiuma, produced by spontaneous contraction of the protoplasm.

FIG. 13.-Representation of a coloured blood-corpuscle of the Amphiuma, the membraneous layer of which has burst; a, colourless protoplasm exposed to view; b, membraneous layer (coloured).

FIG. 14.-Fragment of a coloured blood-corpuscle found in the fresh blood of the Amphiuma.

FIG. 15.-Coloured blood-corpuscle of the Amphiuma, enclosing minute crystals.

FIG. 16.-Another, enclosing vacuoles.

FIG. 17.-Another form of vacuoles.

FIGS. 18, 19, and 20.-Coloured blood-corpuscles of the Amphiuma, treated with water.

FIGS. 21 and 22.-The same, with their nuclei escaping through a rent in the membraneous layer.

FIG. 23.-Nuclei, escaped from the corpuscles, with their contents escaping through a small rent in their enveloping membrane; a, the contents escaping in the form of a small sphere; b, the same, after having been treated with a weak solution of chromic acid; c, the contents escaping in the form of a cone.

FIG. 24. Singular appearance of a coloured blood-corpuscle, treated with water, and subsequently with a weak solution of chromic acid.

FIG. 25.-Coloured blood-corpuscle of Amphiuma, treated with chloroform

vapour.

FIG. 26.-Coloured blood-corpuscle of Amphiuma, treated with chloroform liquid.

acid.

FIG. 27.-Coloured blood-corpuscle of Amphiuma, treated with diluted acetic

VOL. I.

F

A number of years ago, histologists began to deny the existence of an enveloping membrane in the coloured blood-corpuscles, and maintained that the latter consisted of protoplasm only, united with a colouring material, the hæmoglobin. In support of this view, they referred to the bi-concave form of the blood-corpuscles of the Mammalia, and also to the great elasticity which all blood-corpuscles show they possess in assuming almost any form to adapt themselves to the various curvatures and angles of the smallest capillary vessels through which they have to pass; properties which could not be possessed by a vesicular body. The behaviour of the blood-corpuscles towards various reagents, and finally, the impossibility of demonstrating this enveloping membrane, were also adduced to prove its absence.

But while these histologists, by denying the existence of an enveloping membrane, were endeavouring to make the structure of these blood-corpuscles appear of a more simple nature, others, especially in Germany, propounded theories, according to which the construction of these bodies appears quite complex.

In explanation of the various phenomena manifested by the coloured blood-corpuscles, several hypotheses regarding their structure were advanced from time to time. Thus Rollett, denying the existence of the membrane, looks upon them as mainly consisting of a certain stroma, to which they owe their peculiar mechanical properties.* This view is based upon the examination of the colourless remains of the blood-corpuscles of the guinea-pig, horse, and dog, which he deprived of their colouring matter by a peculiar freezing and thawing process. Kuehne, another opponent of the membrane, strongly endorses the view of Rollett.

Another well-known hypothesis, founded upon certain appearances observed on the blood-corpuscles of the Triton, after having been treated with a 2 per cent. solution of boracic acid, is that of Bruecke. According to it, the nucleated blood-corpuscle consists of two parts. The one of these, representing a porous body, of a motionless, very soft, colourless, and transparent substance; the other a living organism, the central portion of which forms the nucleus, filling up the pores of the former, and, with the exception of the colourless nucleus, containing the hæmoglobin. The colourless porous substance, Bruecke named oikoid, the other part zooid. By the contraction of the latter from the former, he tries to explain the occurrence of those appearances observed on the blood-corpuscles of some Amphibia, consisting in the entire or partial retraction of the coloured contents upon the nucleus, and assuming in the latter case the form of a star, as represented by Rollett, on p. 286 of Stricker's Handbuch der Lehre von den Geweben, &c.'

This hypothesis, fanciful as it is, does not explain the pheno* Kuehne, Lehrbuch der Physiolog. Chemie,' p. 190.

menon mentioned, unless the supposed pores represent conical tubes radiating from the centre toward the periphery. But as these have never been seen, of whatever shape they might be, the hypothesis has no foundation of observed facts to rest upon. Nevertheless, it has been received as the true theory by a number of histologists. Even Stricker has adopted this theory with a slight modification.

Hensen, in observing the above-mentioned phenomenon, explained it as caused by the protoplasm of the coloured blood-corpuscles, collected around the nucleus and upon the inner surface of the cell-membrane (which he presupposed to exist), and connected by delicate filaments, radiating from the nucleus to the membrane; the interspaces left between the filaments contain the coloured liquid contents of the cell.

A similar theory, regarding the structure of the coloured bloodcorpuscles of the Frog, has been advanced quite recently by Kollmann. He also presumes the existence of a stroma, formed by a dense network of delicate filaments, extending from the nucleus to the transparent, elastic, enveloping membrane; the interspaces left between the filaments he supposes to contain the hæmoglobin. The characteristic form of the blood-corpuscles he supposes to be only possible by a certain degree of tension in the albuminous filaments of the stroma. An excessive contraction of the filaments is counterbalanced by the hæmoglobin contained in the interspaces. The coloured blood-corpuscles of the Mammalia also, have, according to his view, such a stroma.

Laptschinsky, judging from the effect of different reagents upon the coloured blood-corpuscles of Tritons and Man, supposes them also to consist of two different substances. The one of these, designated by him as the "rest of the blood-corpuscle," is soft and elastic, and assuming mostly a round form, possesses in general all the properties of the so-called "stroma" of these bodies. The second substance becomes under the microscope only visible when, by the effect of different reagents, it is made to swell or precipitate.

Still other hypotheses concerning the structure of the coloured blood-corpuscles and the phenomena which they manifest, have been advanced; but as this article is not intended to present the history of these bodies, I shall forbear mentioning them.

In the course of the numerous investigations into the nature of the coloured blood-corpuscles, especially of the larger ones, containing a nucleus, they have, of course, been treated in various ways-mechanically, chemically, and physically; and there re

*Stricker, Handbuch der Lehre von den Geweben,' p. 295.

† Virchow and Hirsch, Jahresbericht, &c.,' für das Jahr 1874, band i. P. Loc. cit., p. 49.

49.

mains hardly a chemical reagent, liquid or gaseous, to the influence of which they have not been subjected. Heat, moisture, as well as electricity in its different forms, have been applied. The changes occurring in their substance by the action of all these reagents, have been closely watched, and many deductions relating to their structure have been made.

An enumeration of the most important of these experiments will be found in Rollett's article "On the Blood," in Stricker's 'Handbuch der Lehre von den Geweben, &c.,' as well as in other treatises on this subject. But it is certainly strange that while great importance has generally been attached to the various changes occurring in the form of the blood-corpuscles by the action of the particular reagent (which, after all, are only due to a contraction or coagulation of their protoplasm), the double contour of these bodies, the only proof of the presence of a membrane, whether preexistent or artificially produced, is scarcely mentioned, and appears to be generally overlooked or ignored.

In my treatise "On the Origin and Development of the Coloured Blood-corpuscles in Man," published in the February number, 1874, of the "Monthly Microscopical Journal," I made some casual remarks in reference to the fine double contour observed in the human coloured blood-corpuscles after being deprived of their colouring matter by the simple action of that most neutral agent, water. On these stomata, then, as Rollett would call the altered corpuscles in question, a delicate double contour is always detected by close inspection. In directing attention to this double contour, however, I do not presume that its presence indicates a pre-existing true cell-membrane. But, as I have remarked in the above-mentioned article, I do suppose that the coloured blood-corpuscle of Man at maturity undergoes a slight condensation on its surface in the form of a thin layer or pellicle, which, resisting the solvent power of water, finally appears in the form of a double contour. To these conclusions, E. Ray Lankester had also arrived, from a series of observations which he made on the coloured blood-corpuscles, with regard to the action of gases and vapours. The results of these observations were published in the October number, 1871, of the 'Quarterly Journal of Microscopical Science.' "The red blood-corpuscle of the vertebrata," he says in his summary, "is a viscid and at the same time elastic disc, oval or round in outline, its surface being differentiated somewhat from the underlying material, and forming a pellicle or membrane of great tenacity, not distinguishable with the highest powers (whilst the corpuscle is normal and living), and having no pronounced inner limitation."

Lankester, therefore, regards the coloured blood-corpuscle as a viscid and elastic disk, which of course implies that its substance is homogeneous or without structure; he further supposes its surface

to be differentiated from the rest of the body, forming a pellicle or membrane, though of no pronounced inner limitation, and undistinguishable by the microscope. The latter is true with regard to the small, bi-concave blood-corpuscles of the Mammalia; in the large, oval, nucleated corpuscles of the Amphibia, however, the pellicle, as we shall see hereafter, may be readily demonstrated. In the fresh blood of this class of animals we frequently meet with specimens of blood-corpuscles, on which, by a contraction of the protoplasm representing the greater portion of the whole body, the pellicle in question appears separated from the latter, thus manifesting its existence. Lankester himself mentions this fact (p. 368), and gives a representation of the specimens in Fig. 2, a. With due deference to the hypothesis of Bruecke, however, he calls the contracted protoplasm enclosing the nucleus, the zooid, and the separated enveloping pellicle, or membrane, the oikoid. Now, as far as I am able to understand the hypothesis of Bruecke, this oikoid represents a porous, transparent body; and I can therefore not comprehend how Lankester can unite this idea of a porous substance, supposed to embrace the nucleus, and to determine the whole form of the blood-corpuscle, with that of his pellicle or membrane. The hypothesis of Bruecke, ingenious as it is, must eventually prove to be but an unsuccessful attempt to explain certain changes observed to occur in the form of the protoplasm of the coloured blood-corpuscle, which, after all, form the exceptions to the rule, and are such as may be readily explained without ascribing to this body a structure more complicated than can be demonstrated.

A different view from the above is that entertained by Jas. G. Richardson, of Philadelphia. He upholds the old theory, according to which the coloured blood-corpuscles of the Vertebrata are vesicles, each composed of a delicate, colourless, inelastic, porous, and perfectly flexible cell-wall, enclosing a coloured fluid, &c. To this conclusion Richardson arrived, not only by his numerous examinations of the blood-corpuscles of Man and other Mammalia, but also by the study of the corpuscles of the Menobranchus, which, with one exception, are the largest known. An interesting paper, containing these investigations, was republished in the July number, 1871, of the Monthly Microscopical Journal.' In it, he states that he twice succeeded in cutting a corpuscle in two with sharpened needles; and that, on penetrating the vesicle with the edge of the needle, its contents were instantly evacuated, and disappeared at once in the surrounding fluid, while the cell-wall immediately shrunk together, and became twisted upon itself and around the nucleus into a perfectly hyaline particle.

But while Richardson maintains the true cell nature of the coloured blood-corpuscle, and adduced a number of observed facts