View source for Paper - The cortex of the brain in the human embryo during the fourth month with special reference to the so-called Papilla of Retzius

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| [[File:Mark_Hill.jpg|90px|left]] This historic 1907 paper by Streeter described central nervous system neural development. The "papilla of Retzius" was first described by Gustaf Retzius (1842–1919) in  ''Das Menschenhirn''. (1895) Stockholm, one of the first photographic atlases of the brain. Retzius was also a supporter of the "neuron theory" of [[Embryology History - Santiago Ramón y CajalRamón y Cajal]].

See also:
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<pubmed>20979584</pubmed>

:"The fetal cortex consists of a densely packed band of cells in which only the superficial marginal zone (layer 1) is readily distinguished. In fetuses of 16–24 weeks gestation it is not uncommon to see irregularity of the cells of layer 1 and 2 of the cortex with projections of layer 2 cells into the relatively acellular layer 1 or molecular layer (Fig. 1). These are called ‘Papillae of Retzius’ and are commonly seen in the absence of any other abnormality. "

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[[File:George_L._Streeter.jpg|thumb|200px|alt=Embryology History George Streeter|link=Embryology History - George Streeter|George Linius Streeter (1873-1948)]]
=The Cortex of the Brain in the Human Embryo During the Fourth Month with Special Reference to the so-called " Papilla Of Retzius”=


[[Embryology_History_-_George_Streeter|George L. Streeter, M. D.]]

Associate Professor of Neurology at the Wistar Institute.


With 6 Figures.

==Introduction==

The observations reported in the following paper conclusively show that the cortical granulations, or papilla of Retzius, caused by the fungiform arrangement of the cells of the pyramidal layer, and commonly found in human embryos between 11 and 14 cm. long, constitute an abnormal condition, which is produced either by intrauterine or postpartum maceration. It is pointed out that of two human brains of the same age one may have cortical papillae while in the other they may be absent. Furthermore, it is shown in pig brains, where cortical papillae are not normally present, that it is possible by experimental methods to produce a fungiform clumping of the cortical cells that exactly duplicates the condition seen in human brains.


Attention was first called by Retzius, 95, p. 17, to the fact that human brains,' usually of the fourth month and more rarely of the fifth month, possess a fine granulated character perceptible through the smooth surface of the cortex, and in places where the thin superficial layer (Randschicht of His) had been torn off they appear correspondingly granulated or covered with rounded elevations. Microscopical examination of sections of these regions revealed the fact that the granulated character was due to an unequal growth of the pyramidal cell layer, which projected in rounded elevations, the spaces between which were filled in by the superficial or molecular layer, so that the surface of the brain remained smooth. Retzius considered the possibility of this granulation formation being a manifestation of some pathological process, such as is commonly associated with abortion, e. g., syphilis. He was, however, more inclined to believe it a normal condition due to a transitory exuberant growth of the pyramidal cell layer, the surface irregularities caused thereby being smoothed out later by the development of the adjacent layers. Shortly after this Hochstetter, 98, p. 5, briefly refers to the granulations of the pyramidal cell layer described by Retzius. He confirms their presence in poorly preserved brains, and designates this appearance as a decomposition phenomenon, without giving any further evidence.

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| '''Fig. 1.''' Section through the occipital lobe of the brain of a human embryo, 12 cm. long (end of 4th month), showing the irregularities of the pyramidal zone caused by the fungiform clumping of the cortical cells, the so-called papillae of Retzius. Taken from His, 04, Fig. 75.
| '''Fig. 2.''' Section from the brain of a human embryo of about the same age and taken from the same place as shown in Fig. 1. Here the pyramidal zone consists of a compact layer with parallel borders, which, with the exception of three transitory fissures presents a perfectly smooth outer surface, and shows no trace of the Retzius papillae.
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Two years later His, 00, not having noticed the observations of Retzius, described independently the same peculiar granular or wart-like character of the pyramidal layer in embryo brains of the fourth month ; and again in his last work His, 04, he describes at some length this appearance under the title " Die Retziusschen Warzchen." One of his illustrations is reproduced in Fig. 1. Though in his discussion he admits that it is still an open question; yet he is apparently inclined to consider the papilla as normal, and does not hesitate to discard the possibility referred to by Retzius of their being pathological, on the ground that several of his specimens, which showed characteristic cortical papillae, came from healthy individuals who had committed suicide, and the foetuses themselves appeared normal. He also argues that if it were a post-mortem alteration, associated with the swelling of the tissues, then the superficial layer would also present an irregular surface, which is not the case. Neither His nor Retzius accounted for the fact that this phenomenon was not been observed in brains of other animals. With this in mind the writer decided to make the test on some other mammal, fresh embryos of which could be easily obtained at all ages, and Avhere control experiments could be carried out on embryos of the same litter. The pig was selected, and an examination was made of brains of embryo pigs measuring from 10 to 14 cm. long, which is a period that corresponds to and fully covers the time of appearance of cortical papillse in the human brain."" 
It was found that the pyramidal layer in the pig does not exhibit any cortical papillae when carefully preserved ; but has always a smooth, regular surface, the only indentations being those corresponding to the beginning fissures, which make their appearance in specimens between 13 and 13 cm. long. A photograph of a section of a normal brain of a 11.5 cm. pig is shown in Fig, 3.


Having found that cortical papillae are not normally present in the pig, the next step was to see whether they could not be produced artificially, and preferably under conditions which might be probable in case of human material. Two possibilities suggested themselves as etiological factors; in the first place, maceration of the specimen before it was put into the preserving fluid, and secondly imperfect penetration of the preserving fluid. Under maceration we should have to consider both postpartum and intrauterine maceration. The latter might be brought about, for instance, by disease of, or abnormal attachment of the placenta with consequent disturbance of circulation, and perhaps death of the foetus some days before abortion. The second condition, faulty penetration of the fixative, might be present in brains of this size were the fixative not injected through the arteries or the brain coverings not immediately opened up so as to permit the direct action of the fluid on the brain itself.



* The fact that the cortical papillae are usually limited to the fourth month may perhaps be explained as follows: Up to that time the brain wall is relatively thin and uniform in structure, so that deformities then take the form of complete foldings of the wall. In specimens of the fourth month the wall is sufhciently thick to prevent foldings of the entire wall, and expansion and shrinkage express themselves in a readjustment of its constituent parts, some parts being more affected than others. In older specimens such a readjustment is prevented by the development of the cell processes and the supporting framework of neuroglia, resulting in a structure suffiently firm to preserve its form in the fixative, and consequently no more papillse or artificial fissures are found.


The following experiments were carried out with the idea of imitating these two conditions; on the one hand, for obtaining imperfect penetration of the fixing fluid, the brain coverings were not removed until the specimen was ready for embedding, and on the other hand, the maceration was produced either by keeping the specimens dry and exposed to the air long enough for them to macerate in their own fluids before they came into the fixative, or in other cases by putting the brains directly into normal salt solution for varying lengths of time. In human material His found the cortical papillas most marked in material hardened first in formalin and then immersed for several days in Mullers solution. So the same method of fixation was adopted in the experiments, the details of which are as follows :

==A. Maceration Followed by Imperfect Fixation==

A1. Maceration in own fluids (11, 13, and 14 cm. pigs). 
* Embryos left exposed to air, 18 hours. 
* Embryos placed in formalin, 10%, 48 hours. 
* Embryos placed in Muller's solution, 4 days. 
* Washed, brought into alcohol, and then the brain coverings were removed and the brain imbedded and cut in paraffin.


A2. Maceration in normal salt solution (12 cm. pigs). 
* Embryos placed in salt solution, 17 and 48 hours. 
* Embryos placed in formalin, 10%, 48 hours. 
* Embryos placed in Muller's solution, 4 days. 
* Washed, brought into alcohol, and brain removed as above and the brain then sectioned in celloidin and paraffin.


In these specimens, in which the brain coverings were left intact throughout the period of fixation, no cortical papillae were found. Embryos of different sizes were tried (Al) for the purpose of covering the whole period favorable to the formation of the papilke. The poor preservation of the tissues manifested itself by a varying degree of fragmentation of the sections, particularly of the deeper parts. The sections presented a shredded appearance which varied from minute forking clefts between small clumps of cells and between fiber bundles, up to large irregular cracks splitting the different layers of the brain walk This condition was found both in material that was cut in paraffin and in that cut in celloidin, but it was more marked in specimens that had been macerated in salt solution 17 hours, and still more marked in those macerated 48 hours. Otherwise the general topography of the sections and the arrangement of the layers was fairly well preserved. The minute clefts between the cells of the pyramidal layer gave a slightly ragged appearance to the surface of that layer, but it was nothing that approached the fungiform arrangement seen in the Retziiis papiHs. As can be seen in normal specimens at this time, the pyramidal layer is split by a line of scanty nuclei into a more superficial thicker subdivision, the pyramidal cells proper, and a deeper subdivision which is to form the layer of polygonal cells. This stratification was preserved in the experimental material. Another feature of importance was the absence of the so-called transitory fissures.


[[File:Streeter1907 fig03.jpg|400px]]

'''Fig. 3.''' Section from a well-preserved brain of a pig embryo. 11.5 cm. long. This section shows that normally, in the pig brain of this age, the pyramidal zone presents a uniformly smooth outer surface. This brain, while still warm, was preserved in a chrome-acetic mixture.

[[File:Streeter1907 fig04.jpg|400px]]

'''Fig. 4.''' Section from a macerated brain of a pig embryo, 11.5 cm. long. The brain was kept in normal salt solution 48 hours and then preserved in formalin followed by Muller's solution. The section shows distinct fungiform clumping of the cortical cells and characteristic Retzius papillae. The same specimen is shown under higher power in Fig. 6.

==B. Maceration Followed by Good Fixation==

B1. Maceration in normal salt solution (11.5 cm. pigs) 
(a) Fresh brain placed in salt solution, 28 hours. Hardened in chrome-acetic solution, 48 hours. Washed, dehydrated and sectioned in celloidin.

(b) (Figs. 4 and 6) Fresh brain placed in salt solution, 48 hours. Hardened in formalin 10%, 24 hours. Secondary fixation in Muller's solution, 4 days. Washed, dehydrated and sectioned in celloidin, 

B2. Maceration in its own fluids (11.5 cm. pig) see Fig. 5. Embryo left exposed to air, 48 hours. Brain removed and kept in formalin, 48 hours. Secondary fixation in Muller's solution, 4 days. Washed, dehydrated and sectioned in celloidin.


The sections of the specimens macerated in salt solution (Bl, a and b) show fairly good preservation of the deeper lying parts, there is almost no shredding of the tissue like that seen in the specimens in which the penetration of the fixing fluids was hindered by the brain coverings. The pyramidal layer of the cortex, however, is found to be thrown into irregular folds, accompanied by a fungiform clumping of its constituent cells. This appearance is present in both specimens, but is more marked in the specimen (&) macerated 48 hours. A section of this was photographed and is reproduced in Fig. 6. The resemblance is close to the description given by His and Retzius of the cortical papillae in the human embryo. It has the same smooth-surfaced superficial layer, which dips down between the papillae of the subjacent pyramidal layer. In some places these incisures cut off small irregular islands of pyramidal cells. The inner surface of the pyramidal layer does not have these sharp notches, but runs across the section in an irregular wavy ill-defined line. In addition to the fungiform clumping of the cortical cells, in some of the sections the so-called transitory fissures are found. These dip sharply inward and invade in some cases more than one-third of the thickness of the brain wall. In the formation of these, the superficial layer is partially folded in with a corresponding cleft on the surface of the brain, which is not the case with the cortical papillae. It may be noted that the artificial character of transitory fissures has been well established by Hochstetter, 98, and Mall, 03, the latter having examined over fifty embryos and found that according to the effect of various dissociating influences he could obtain maceration in all stages, from simple folding of the brain wall up to conversion of the entire central nervous system into a pulpy mass. Evidently cortical papillae and transitory fissures, though differing in character, have a similar etiology; as in the above experiment we have both, artificially produced in the same brain under known conditions. The interesting fact should be noted that though the two formations may occur in the same brain, and may closely adjoin each other, yet they do not occur together at the same place ; that is to say, one does not find a fungiform grouping of the cells that lie in the cleft of a transitory fissure. Either process seems sufficient to satisfy the space demand.

[[File:Streeter1907 fig05.jpg|400px]]

'''Fig. 5.''' Section of the brain of a pig embryo, 11.5 cm. long. The embryo was left exposed to the air, and the brain allowed to macerate in its own fluid 48 hours. The brain was then removed and preserved in formalin followed by Muller's solution. In this section the fungiform clumping involves only the outer part of the pyramidal zone, and in this respect closely resembles the condition seen in Fig. 1.

[[File:Streeter1907 fig06.jpg|400px]]

''Fig. 6.'' Section of same specimen shown in Fig. 4. The maceration here is more advanced than that seen in Fig. 5. The fungiform clumping involves the whole thickness of the pyramidal zone, both the inner and outer surfaces of which are thrown into coarse irregular folds.
 
Sections from the specimen macerated in its own fluids (B2), see Eig. 5, differ from those macerated directly in salt solution in that the fungiform arrangement of the cortical cells involves only the more superficial part of the pyramidal layer. Instead of foldings of the whole layer, such as is seen in Fig. 6, we have here only a granulated or fungiform surface ; and this duplicates almost exactly the condition found in the embryo PI of His, which he pictures in Figs. 75, 77, 98 and 99. It shows that it is possible by different methods of maceration to produce experimentally typical cortical papillae in brains where they are not normally present.


==Conclusion==


The comparison of Figs.1 and 3, one with, and one without cortical papillae, suggests the probability of the abnormal character of the papillae. One could still perhaps raise the objection that they may be normal, but very transitory, and that the two sections do not quite represent the same stage of development, so that in Fig. 2 the papillge have either already disappeared or have not yet developed. This objection, however, can no longer be considered in face of the fact that in pigs, where one is able to secure specimens in exactly the same stage of development, it is possible, as has been shown above, to produce the papillae by means of maceration, and furthermore to control their size and character by varying the degree and method of maceration.


From the experience derived from the above experiments, as regards conditions which predispose to artificial fissures of the cortex and deformities of its constituent cell layers, it becomes evident that embryo brains, which are intended for general morphological study, should, up until the time of completion of the principal fissures, be hardened in situ without disturbing the brain coverings. If the brain of a human embryo fresh from the uterus is uncovered or completely removed, and then immediately immersed in formalin or other fixative, it will not necessarily be free from abnormal fissures, etc. The framework of the brain wall up to that time is by no means firm, and it must be also remembered that it may already have been softened by maceration in the uterus. Thus in such a case, and much more so in the embryos that do not reach the hardening fluid so promptly, it is essential that the ))rain coverings should be left intact, that they may serve as a support to the brain during the process of fixation. Imperfect penetration of the preserving fluid is to be obviated by injecting it through the blood-vascular system.

==Literature==

His, W., 00.  -  Developpement de la substance grise de I'ecorce cerebrale. XI He Congres international de Medecine. Paris, 1900.


04.  -  Die Entwickelung des menschlichen Gehirns. Leipzig, 1904.


HocHSTETTER, F., 98.  -  BeitragB zur Entwickelungsgeschichte des Gehirns. Bibliotheca Medica, A. Heft 2., Stuttgart, 1898.


Mall. F. P., 03. -  On the transitory or artificial fissures of the human cerebrum. Amer. Jour, of Anat., Vol. II.


Retzius, G., 95.  -  Das Menschenhirn. Stockholm, 1895.

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