THE 
CEREBELLAR CORTEX OF THE DOG. 
By HENRY J. BERKLEY, M.D. 
From the Johns Hopkins Hospital Reports, Vol. Ill, Nos. 4, 5 and 6, Baltimore, 1893. THE CEREBELLAR CORTEX OF THE DOG. THE CEREBELLAR CORTEX OF THE DOG. 
HENRY J. BERKLEY, M. D 
The investigation was undertaken to determine if the use of some 
of the newer methods of hardening and staining, in connection with 
the old, together with a collective view of a large number of sections 
and teasings differently treated, would not lighten to some extent 
the obscurity now existing concerning the fine anatomy of this, the 
most complex organ of the central nervous system, or of the entire 
body. 
The limits of the paper will not enable me to more than touch 
occasionally upon the works of others (Deiters, Koschewnikoff, Had- 
lich, Schwalbe, Dennissenko, Beevor, Obersteiner, etc.), nor to enter 
at length upon the discussion of disputed points: it is simply a 
resume of what I have myself seen in the course of a series of 
examinations. 
Dogs, cats and pigeons were at first used, but the cerebellum of 
the first-named animal presented so many points of superiority over 
the others, both in the tissue itself, and the ease and rapidity with 
which it could be removed from the cranium, that the others were 
abandoned for it. As a matter of fact, it was found possible to 
chloroform, remove the organ, and have the pieces of the practically 
living tissue in the hardening media within five minutes after the 
commencement of the narcosis. All the animals used were young, 
but full grown. 
The following methods of hardening and staining were employed : 
1. a. Flemming’s fixation fluid, into which very small pieces of 
the cerebellar tissue were placed for from twenty-four to thirty-six 
hours at the room temperature of from 20° to 25° C. 
b. The same process, except that the fluid was heated to 35° C. 
before the tissue was placed in it, and kept at that temperature for 24 
hours in the warm chamber. Absolute alcohol after hardening for 
both series. The sections from a were the best. 
The sections from this series were stained with safranin, generally 
for twenty-four hours, occasionally for several days; with carbolic- 196 
Henry J. Berkley. 
acid fuchsin; and with ray modification of Weigert’s hematoxylin.1 
The safranin and osraic-hematoxylin preparations proved the most 
satisfactory of all the preparations. Unstained sections were also 
examined with comparatively good results. 
2. a. Muller’s fluid employed in the same way as the Flemming, 
namely, small pieces being dropped into the cold fluid and allowed to 
remain in it for six weeks, the fluid being frequently renewed in 
the meanwhile, and carefully kept from the light and air. 
6. Cubes not more than half a centimetre square were placed in a 
large jar of fluid heated to 35° C, and the temperature maintained at 
that degree for ten days, the fluid being changed every second day. 
After the tenth day the jar was removed from the hot-air oven, and 
afterwards, as in the first method, the cubes were dehydrated in 
alcohol without previous washing in water. 
The stains used with the Muller’s fluid preparations were Wei- 
gert’s hematoxylin, aqueous fuchsin, nigrosine, Weigert’s saure 
fuchsin, borax carmine, alum-hematoxylin, eosin, and the carminate 
of soda staining in toto after Schultze. 
The copper-hematoxylin, fuchsin, hematoxylin, and sodium 
carminate were the most useful. Combinations were frequently 
employed. 
3. Disassociated preparations were also made from the fresh 
tissue, and after maceration in dilute Muller’s fluid and osmic acid 
solutions, with fair results, principally to confirm certain points in 
sections. Some fresh preparations were stained in aqueous fuchsin. 
4. The sublimate method with the quadruple stain of Gaule; also 
sections from sublimate stained with borax carmine. Neither pre- 
sented any advantages over other methods. 
5. Hardening in absolute alcohol, and after treatment according 
to the method of Nissl, proved extremely satisfactory for the cellular 
elements and glia tissue. Carmine tingeing also gave some very 
good specimens. 
6. The sublimate method of Golgi, also as modified by Pal, pro- 
duced some useful slides, though the terminal protoplasmic filaments 
of the Purkinje cells were brought out more satisfactorily by carbolic 
fuchsin with Flemming, and a modification of the Weigert copper 
stain, in which the metal is finely deposited upon the protoplasm, 
'See Johns Hopkins Hospital Bulletin, No. XII, 1891. The Cerebellar Cortex of the Dog. 
197 
the medullated tubes retaining their deep blue, while the cellular 
substance assumes a brown tint. 
Imbedding wras always done in celloidin, except in one or two 
instances; very often directly from the absolute alcohol into the 
ether-alcohol-celloidin mixture, which was found to answer perfectly. 
Sections were cut of varying thickness, from one down to less than 
half a space on the Schanze microtome. These sections were not 
confined to parallel and vertical cuts, but were made at a large num- 
ber of angles to the surface, acute and oblique; and from these, some 
important items concerning the direction of nerve and other fibres 
are deduced. Serial sections were only made in a single instance, to 
determine the space between the Purkinje cell rows. 
The Central Core and Medullated Fibres. 
The central core of the lobules as it passes off from the deeper 
white masses into the leaflets presents no difference in the size or 
contour of its fibres from the ordinary structure of white medullated 
tissue. Now and then, scattered multipolar nerve cells are seen in 
it at the base of the folia; a few larger than those of the great cell 
layer of the cortex, and much resembling them in contour. These 
cells are isolated offshoots from the individual groups that always 
lie lower down in the white substance. 
Scattered among the fibres are nuclei in about the same proportion 
as in the cerebral medullary matter. These nuclei are mainly oval, 
sometimes round; with Nissl’s method very finely grained, with two 
or three larger particles of chromatin scattered irregularly through 
the nuclear substance. The oval nuclei show a small amount of 
protoplasm capping the end, and from this, delicate unstained 
fibrillae, extending between the medullated fibres, may be seen. 
Ganglion cells, isomeric with the Purkinje, are met with, though but 
rarely, along the border of the granule layer. 
As the core is traced toward the apex of the leaflet, fibres are seen 
to pass off from it at an oblique angle into the red-brown layer, until 
at the apex, at a distance of from two to three millimetres beneath 
the large cell layer, it spreads out like the sticks of a fan, with a 
somewhat regular arrangement. 198 
Henry J. Berkley. 
The fibres may be easily separated into several classes, (1) a series 
of apparently unbranched, non-varicose, straight, medullated tubes, 
passing directly from the sides and apex of the core to the vicinity of 
the Purkinje cells. Some pass straight, others bend as they approach 
the “ limitans interna”1 to reach their individual cell. In sections 
perfectly plane to the surface, their connection with the Purkinje cells 
may often be demonstrated. As they enter the cell’s capsule at its 
inferior pole, the medullated sheath is lost, or rather becomes 
directly continuous with the capsule of the cell, and is indistinguish- 
able from it (Fig. 3). Rarely the fibre has the appearance of 
becoming non-medullated just before the capsule is reached, but such 
appearances are exceptional, and may be deceptive from the out- 
spreading of the glia fibres from the capsule. 
The axis cylinder is lost immediately after entering the protoplasm 
of the inferior pole, so far as I have been able to discover, but the 
capsule hides the true relation to some extent. Disassociated prep- 
arations from dilute Muller’s fluid, unstained, or tinged with fuchsin, 
also show distinctly the axis cylinder entering the protoplasm, and 
rarely, a light striation may be discovered. 
The second class also pass to or from the central core, generally by 
one of the so-called eosin cells of Dennissenko, and after continuing 
by it, branch indefinitely between the groups of granule cells ; though 
through the densest of these groups the coarser fibres seldom seem to 
run; then continue by other eosin cells, and so on, forming an open 
network of a very complicated arrangement. 
I have not been able to determine positively if all these fibres pass 
in close relation to an eosin cell, as they go to or from the core, 
before branching, but am of the opinion that some connect with other 
fibres before touching any of these bodies. 
The fibres composing the network vary in size considerably, being 
seldom so coarse as the fibres of the first class, then passing in various 
gradations to the finest kind of medullated fibres, as fine as any 
demonstrable in the cerebral cortex. Even with the finest of them 
Weigert’s reaction never fails, and I do not regard any of the fibres of 
the inter-granular plexus as being of non-medullated character. 
Indeed, very few non-medullated fibres can be determined in this 
zone, and all that are present are probably in direct relation with the 
1 See page 203. The Cerebellar Cortex of the Dog. 
199 
scattered ganglion cells in this layer. At the point of juncture 
between fibres there is always a slight thickening of the medullary 
covering ; generally the fibres are somewhat varicose. 
Under the “limitans interna” branches shoot out from the plexus 
in numbers, and proceed vertically, or at a right angle into the mole- 
cular layer, where they rapidly attenuate, lose their medullated char- 
acter, and for the most part are lost in the inferior and middle 
thirds, mainly in the neighborhood of certain small nerve cells lying 
in this region; while a minimum number continue toward the peri- 
phery, also losing their medulla and becoming reduced to the finest 
threads, when they can no longer be traced. Few continue beyond 
the middle of the layer. Numbers run for considerable distances in 
the band of tangential fibres, and then turn upward into the outermost 
layer. 
The third class of medullated fibres are not very numerous. They 
proceed like the first class directly from the core through the granule 
layer, pass between the Purkinje cell bodies, then slanting consider- 
ably in their course, traverse a part of the barren zone, rapidly 
becoming attenuated and non-medullated, and disappear in the 
neighborhood of the before-mentioned small cells in fine terminations. 
Likewise these apparently do not branch. 
Associating Fibres.—To the fibrse arciformes connecting the folia, 
no reference is made in this paper. Within each leaflet but one 
system that bears any approach to an association can be distinguished. 
This lies about and covers the capsule of the Purkinje cells with a 
thin layer of fibres, with a general arrangement and direction similar 
to that of the great cell row, but also around each cell’s capsule a few 
fibres are wound like loose threads upon a ball. There is an apparent 
variety in the quantity of the fibres in each leaflet, but they are usually 
most numerous where the Purkinje cells are thickly set. There is also a 
difference in the breadth of the band in different leaflets. It never 
extends much beyond the inner limit of the “limitans interna on the 
other hand, its circular fibres may be met with as high as the central 
part of the molecular layer, but widely scattered. Generally its margin 
is within the outer limit of the inner third of this layer. Allowances 
are to be made for possible differences- in staining. The fibres com- 
posing it are medium-sized medullated tubes, varying slightly in 
fineness, the medulla is double-contoured and stains easily blue-black 
with Weigert’s hematoxylin. 200 
Henry J. Berkley. 
In the outer zones of the molecular layer a few scattered medul- 
lated tubes may be seen, with a direction parallel or oblique to the 
pia; but they are insufficient in number to form any approach to a 
system. 
Crossing the central core at right angles to the longitudinal fibres, 
medullated tubes may be found in inconsiderable numbers, running 
from the anastomosing fibres of one side to those of the other half 
of the granule zone. 
The continuance of the straight, apparently unbranched fibre into 
the inferior pole of the Purkinje cell has been moderately definitely 
made out before, notably by Beevor, and the only uncertainty seems 
to be as to its manner of entering the pole. Fig. 3, drawn with the 
aid of a Zeiss immersion system, shows as nearly as possible the 
relation, though as before remarked the capsule hides it very often. 
It is simply an hypothesis, but I am inclined to regard them as 
efferent fibres that run downwards to the cord, or more probably to 
nuclei of the medulla oblongata, their nerve impulses being continued 
further through a second set of cells; while those fibres that first lose 
their medullary sheath in the upper part of the inferior third of the 
molecular zone may be the afferent from the spinal cord. Many of 
them terminate near the small ganglion cells of the molecular layer, 
and probably have a different function from those coming from the 
Purkinje bodies. It is possible that the fibres of the third system 
may be in connection with those of the nerve cells of the molecular 
layer also, in which case they may also be considered as afferent. A 
portion of the fibres which arise from the nerve cells of the barren 
and granular zones may very possibly convey impulses to the cere- 
brum. The disappearance of a large part of the fibres passing from 
the anastomosing plexus into the barren zone, in close proximity to 
certain ganglionic bodies, leads to a suspicion of a direct relation 
between the two, and it is more than possible that the great majority 
of the fibres of the granular layer are centripetal-coursing. 
It is difficult to understand the reason why an anastomosing net- 
work should have an association system at its outer edge, yet the only 
fibres that run into it in considerable numbers, and lie parallel with 
the band for any distance, belong to the plexus and may be traced 
directly into it. The Cerebellar Cortex of the Dog. 
201 
As the band is generally best developed where the Purkinje cells 
are thickly set, and as all the fibres passing to and from the mole- 
cular layer pass through it, it may have the function of uniting, or 
rather transmitting the nervous impulses derived from the various 
kinds of cells in the cortex into one common channel, should neces- 
sity arise therefor. 
The Granule Zone.—The limits of this layer are the least defined 
of any in the cerebellar cortex. Narrow at the base of the leaflets, it 
gradually spreads out to a breadth several times exceeding that of 
the inferior part. Likewise its round nuclei penetrate everywhere 
into the edges of the central white core, and to some extent along the 
borders of the molecular layer, though here the round nuclei are 
replaced in large measure by cells of another description. 
Within these limits six different varieties of cellular bodies, exclu- 
sive of those appertaining to the blood-vessels, may be distinguished. 
The most numerous are the variously named, round, granule, or 
hematoxylin cells (Figs. 1, 2, 6), which were the distinguishing 
feature of the layer with the old carmine preparations, and with the 
addition of a small ring of protoplasm around each nucleus, make 
up more than half the substance of the zone. These round nuclei are 
a little smaller than those of the neuroglia in medullated tissues, but 
are unlike them in taking up much more of the hematoxylin, safranin, 
or magenta dyes, hence stand out more prominently in the field of the 
microscope. They are arranged with a certain definite grouping 
(Fig. 2), which is most pronounced at the apex of the leaflet, and 
shows spaces between each group, which are filled by the eosin 
cells, glia fibrillse, and medullated tubes. The nuclei are always 
perfectly round, contain numerous small molecules deeply tinged; 
the smallest being arranged along the periphery, the larger somewhat 
more centrally, with always a single one of a greater size than the 
others, giving to a limited extent the appearance of a nucleolus. 
Around the nucleus is a diminutive ring of an apparently proto- 
plasmic substance, easiest seen with the nigrosine stain, but at best 
difficult to distinguish in sections stained in any manner, as it 
generally remains untinged. In teased preparations, and in sections 
separated by pressure on the cover glass, it is, however, more clearly 
brought out. Threads extending from this protoplasmic ring are 
demonstrated with extreme difficulty, and I have only succeeded in a 202 
Henry J. Berkley. 
few instances in convincing myself that they are present, then only in 
teased slides. 
Widely but sparingly distributed through this layer are round, 
sometimes slightly oval nuclei, with a nuclear substance that does 
not take up any of the aniline dyes, but only shows a few minute 
molecules clustered along the margin, and an invariably distinct and 
well-tinged nucleolus lying in the centre (Figs. 2, 5). These clear 
nuclei are present in the spaees between the granule clumps, and are 
about double the size of the granules. I have never succeeded in 
isolating them with any protoplasm attached, nor distinguished 
fibrillse proceeding from them. They are probably glia-tissue 
elements, having their prototypes among similar nuclei in the medul- 
lated and molecular regions. 
The undoubted glia nuclei are separated into three different varie- 
ties, distinguished chiefly by their form and some difference in size. 
In the central and deeper regions of the zone they are infrequent, 
lying chiefly near the white fibres, for which their threads prob- 
ably serve as supports. Close to the Purkinje cells they constantly 
increase in numbers, while the granule nuclei as constantly diminish, 
until between the lower half of the bodies of these cells (which is, 
however, properly in the limits of the molecular zone) but few of 
any kind of nuclei other than glia can be found. 
Of the three varieties, a very small round nucleus, about the size 
of the granule nuclei, are to be seen in deeper parts, together with a 
few of an elongated oval form (Fig. 5), both lightly tinged and 
holding a few chromatin particles. In the same region may be seen 
nuclei tipped with a hood of protoplasm (Fig. 5), which sends off* 
from its apex one prominent stem. This protoplasmic hood is 
apparently separate from the nucleus. 
Along the margin of the “ limitans interna ” the round and pyra- 
midal cells assume the importance of a double or triple, though 
irregular row, mainly with their broader bases directed inward; and 
from the latter nuclei come off* very numerous fibrils ascending into 
the molecular layer (Fig. 1). At rather definite intervals a stronger 
thread ascends to the sub-pial limit, and is attached in the thickened 
glia on the periphery. In the central regions the glia fibrils, like 
their nuclei, are but poorly developed, the granule cell groups being 
so closely packed that with the other elements, inclusive of the The Cerebellar Cortex of the Dog. 
203 
capillaries between them, there is but little room for any accessory 
fibre development. 
But the aspect along the outer edge is widely different, the fibrillse 
from the very numerous glia cells expand to a vast open network, 
wrapping the Purkinje bodies in a thick envelope, twisting and 
crossing between the nerve tubes, vessel plexus, and other cells in 
all directions. The name “ limitans interna ” has been given to this 
network of the neuroglia by Beevor. 
Eosin Bodies.—The so-called eosin cells of Dennissenko form 
apparently a unicum in the histology of the central nervous system. 
They consist of spindle, triangular (which largely predominate, and 
show a triangular aspect both on vertical and horizontal section), 
and irregularly quadrate bodies; confined entirely between the 
extreme inner limit of the “ limitans interna ” and the central core, 
into which they press to some extent, depending upon the density 
or rarefaction of the nerve fibres. They are somewhat more numer- 
ous toward the apex and central regions of the granule zone, but 
present few modifications in size or general appearance in its differ- 
ent portions. For actual examination they stain only with the 
Flemming-copper-hematoxylin, and the Weigert so modified to pro- 
duce a copper precipitate. Eosin, nigrosine, and fuchsin also tinge 
them, but with the exception of the last the definition is far from 
perfect. Something may be done with teased preparations, especially 
after they have been some time in dilute osmic acid solution. 
The eosin bodies are arranged in irregular collections, with rows 
or masses of granule cells separating them; and wherever the nerve 
fibres of the anastomosing plexus lie thickest, there also the eosin 
cells are best developed. Their size varies from about 13x10//up 
to 18 x21/i for the triangular; 13x15/* to 33x19// for the largest 
and smallest spindle, and about 9x8// for the irregularly shaped 
ones. The thickness is about the same as the breadth. The cell 
body is formed in large proportion of coarsely granular particles, 
with here and there irregularly placed larger ones (Figs. 1, 4), both 
round and rod-like in shape, all imbedded in a small amount of a 
homogeneous matrix, which can only be tinged effectually after treat- 
ment with osmic acid (carbol-fuchsin, and Flemming-hematoxylin). 
No nucleus or necleolus is anywhere visible in any of them, the 
closest approach to it being in unstained or safranin preparations, 204 
Henry J. Berkley. 
where there occasionally appears a vacuolated appearance toward 
the centre of the body, which resolves, after osmic acid treatment, into 
one or more of the largest granules lying close together. 
The relation of the anastomosing nerve fibres to these granular 
bodies is peculiar and interesting. With saure and carbolic fuchsin 
the impression is given that the nerve tube enters the body, loses its 
medullated sheath, and regains it beyond; but this is not confirmed 
by either the Flemming or Weigert hematoxylin stains, nor by the 
safranin method of Adamkiewitz. The nerve fibres are now seen to 
run in close approximation to the granular body (Fig. 4), over it, or 
along its margin, but do not directly enter. Occasionally there is 
the appearance as if a few of the finer granules were thrown over the 
medullated sheath, but this is rare, and may be deceptive. 
Numbers of all, but especially the triangular forms, are sur- 
rounded on two or even on all three sides by the medullated tubes, 
so as to present the appearance of a blue-black triangle inclosing a 
brown centre (copper precipitate preparation). The fibres after pass- 
ing beyond the body again join the network and proceed to branch 
indefinitely. Nowhere can any connection between the axis cylinder 
and the granular cells be demonstrated, nor does the medullated 
sheath lose its contour. Fibres also cross the long and transverse 
diameter of these bodies in all directions, but apparently only touch- 
ing their surface, sometimes even branching as they reach the mar- 
gin, then sending a double fibre over the surface to finally disappear 
in the network. 
Very fine-grained processes can be seen extending from the poles 
of the cells, in sharp contrast by their brown color with the medul- 
lated fibres, their terminations disappearing between the rows of the 
hematoxylin nuclei. Besides these rami there may be seen exceed- 
ingly fine threads coming apparently from the body, which, on first 
examination, also look like extensions, but with immersion systems 
they are seen to have an exceedingly thin layer of blue medullated 
substance around a darker centre, and when they can be traced any 
distance join fibres of the branching system. Accordingly, they 
probably rise from layers beneath the section, to the body, then pass 
on a level with it to connect with other fibres of the same system. 
Around isolated granular cells a number of unstained, exceedingly 
fine filaments may be distinguished by their refraction, but these The Cerebellar Cortex of the Dog. 
205 
are probably derived from the neuroglia, which is better developed 
in their neighborhood than elsewhere in the granule zone. 
Throughout this zone a considerable number of multipolar, 
bipolar, and pyramidal-shaped nerve cells are to be found scattered 
here and there. They have well developed nuclei, nucleoli, and 
processes, the last soon becoming lost among the granule cells. 
There is also an extremely rare, very large, irregularly-shaped nerve 
cell, with an excessively large, clear, oval nucleus, with few chromatin 
particles in it and a bright small nucleolus. The protoplasm and 
extensions are with any method of staining very ill defined (Fig. 6). 
There is no discoverable relation of any of these cells to nerve fibres, 
though, of course, it must exist. 
The question whether the granules are the nuclei of nerve or glia 
cells has been for many years a much combated point. My 
researches have led me to place them, like certain other small cells 
in the posterior horns of the spinal cord, among nerve formations, 
and upon the following grounds : In certain diseases of the cerebel- 
lum they, in common with the ordinary fragile nerve elements, 
atrophy, and even disappear entirely, while the neuroglia cells 
usually remain and are increased in quantity. In a recent case of 
the so-called gliosis of the cerebellum, a portion of the organ was 
hardened in Flemming, and stained in safranin and osmium-copper- 
hematoxylin. 
The whole granule layer viewed with a low power appeared less 
dense than usual; when examined with an immersion system, the 
granules were seen to be wider apart, looked more coarsely grained 
than normal, while among the groups were ill-stained fragments of 
the cells, apparently undergoing a disintegrative process. There was, 
moreover, a difference in the absorption of the dye by the nuclear 
substance generally. A distinct disappearance of the medullated tubes 
and eosin cells of the layer was also quite evident; the increased 
space being filled in by an abundant growth of rather coarse fibres, 
while the nuclei of the neuroglia stood out quite prominently. In 
complete sclerotic degeneration of a cerebellar leaflet or lobule from 
arterial disease, the granules, in common with the more prominent 
nerve elements, completely disappear. 206 
Henry J. Berkley. 
The presence of an exceedingly small amount of ordinary neuroglia 
tissues in the granule layer has been previously mentioned by Ober- 
steiner and others, who give no detailed description. This small 
proportion in the central parts of the layer has very recently been 
demonstrated by Weigert, by means of his new method of staining 
the neuroglia tissue solely.1 
The eosin cells, the most peculiar and striking feature of this 
layer, are not the least explicable element of this most intricate 
organ. They possess most of the attributes of a cell—and a nerve 
cell at that—protoplasm coarsely granular, prolongations, etc., but 
the nucleus is wanting, or at least cannot be demonstrated by any 
known method of staining, or be discovered in the unstained cell; 
yet if they are nerve cells it is a necessity that they have a nucleus, 
as no exception is known in the animal body. It is possible that 
the nucleus is very small, and may be hidden in the thick-set coarse 
granules that cover and overshadow it. If nerve cells they be, they 
are probably subsidiary to cells of higher function, the Purkinje for 
instance, and possess a connection with the fibres of the anastomosing 
plexus through their fine protoplasmic processes, that are utterly 
impossible to trace for any considerable distance from the cell body, 
through the thickly intervening granules. 
Their intimate relation to the medullated fibres is singular, and 
does not exist elsewhere in the nervous system. There are places in 
the layer where medullated fibres are scattered, in which there are no 
eosin cells, but where the eosin bodies are numerous the medul- 
lated fibres multiply in numbers, seemingly not on account of an 
origin from these bodies, but because they seem to be placed along 
their paths. 
Why should they enter into such intimate contact with these cells, 
curving along their rounded borders, or passing in close contact over 
them, unless they received some impulse from them? Yet the medul- 
lated sheath certainly continues uninterruptedly by the bodies; and 
the axis cylinder, so far as can be determined, does not divert any 
fibrillse at the point of closest relation, nor is there any thickening 
of the medullary covering, as is seen elsewhere when the fibres 
branch in the plexus. Their complete atrophy in common with the 
undoubted nerve elements helps to classify them among the same 
structures. 
'Anat. Anzeiger, No. 19, 1890. The Cerebellar Cortex of the Dog. 
207 
The Great Cell Row and Molecular Layer. 
The Purkinje cells form the most prominent connecting link 
between the granule and barren layers of the cerebellum. They are 
flask-shaped protoplasmic bodies, with their central prolongations 
passing through the “limitans interna,” to be united with one of the 
radiating coarse medullated fibres that enter the central core. Best 
developed at the apex of the leaflet, they become somewhat less 
numerous in the depths of the furrows. In size they are about the 
same as in man, measuring 38// in their long diameter, 25// in their 
transverse, and from 20 to 23// in thickness. The rows of cells are 
not perfectly parallel, and in cross sections are arranged a little 
irregularly in each line. Nearly surrounding them on all sides are 
the numerous glia cells, from which their capsule of the finest felt- 
work of fibres is derived. This enveloping is very clearly and 
distinctly seen in Flemming and copper-precipitate preparations. 
Where the medullated fibre enters the inferior portion of the central 
pole, its sheath immediately enters into and becomes a part of it, or 
at least is not distinguishable from it, staining the same hue, and 
the axis cylinder is lost in the cellular protoplasm, which for some 
distance is fibrillated. At this point of entrance the capsule is 
thickest and more dense than at the sides and apex (Fig. 3). The 
capsule extends rather further down on the central pole than is 
drawn in the figure. This capsule is continued upward on the chief 
prolongations, gradually diminishing in thickness, until it becomes 
over the medium-sized processes a microscopically structureless 
membrane, supported by, and in close relation with the neuroglia 
tissue and perivascular spaces of the smallest vessels in the mole- 
cular layer, and is doubtless a portion of the glia formation.1 This 
enveloping of the peripheral prolongations is distinct in chrome and 
Flemming preparations (carbol-fuchsin, safranin, also in unstained 
sections), less easily demonstrated in alcohol ones. 
The encapsuling of the Purkinje cell gives a lymph space which, 
for the barren layer taken as a whole, is of stupendous extent. Hori- 
zontal sections of the cortex in the inferior third of the zone also 
show the cell branches with a clear ring around them (Miiller-car- 
1 The finer portion of the capsule may possibly be derived from the hyaline sheath 
of the vessels ; the thicker portion is certainly made up from fibrillae. 208 
Henry J. Berkley. 
mine-nigrosine). Mixed with the glia fibres surrounding the body 
of the cell are numbers of very fine naked axis-cylinder threads, 
apparently coming from the molecular layer to reach the cell. 
The processes that extend peripheral wards give a variety to the 
appearance of these bodies. Some, and that the majority, have a 
single thick process, which usually leaves the body perpendicularly 
to the periphery; while others have a thick process that almost imme- 
diately divides; and there is a third variety in which the arms leave 
the cell at opposite angles (Fig. 1). 
With unstained, and with most of the usual methods of tingeing, 
the body of the cell appears finely granulated, with a certain concen- 
tric arrangement, which gradually disappears at the upper pole, 
beyond which the prolongation usually shows a light striated appear- 
ance, which grows less and less distinct, disappearing entirely in the 
finer branches. The inferior pole of the cell has also sometimes a 
striated arrangement. NissPs stain gives a number of coarse gran- 
ules among the finer ones of the protoplasm. The cell body never 
contains any pigment deposit. 
The nuclei are invariably clear, round or slightly oval, and hold a 
number of round bright-colored particles that with Flemming are 
often rod or threadlike, giving something of a radiation around the 
nucleolus. This nucleolus is rounded, very often slightly irregular, 
colors brightly, and is finely grained. 
From the protoplasmic processes of the preceding paragraph thick 
branches arise, that again and again subdivide, almost invariably at 
right angles to the periphery, until they become filaments of tenuity, 
and form a feltwork inconceivable in its magnitude and fineness. 
From the first branches that come off from the main stems at 
an angle of about 180° to the surface, filaments pass upward and 
downwards, the latter filling the space to some extent between the 
thick rectangular processes, and the small nerve, and pear-shaped 
glia nuclei. The former pass upward, in distance commensurate 
with their thickness, until they become too fine to be followed. Like 
the upper branches of a tree, denser and denser does the feltwork 
become as the periphery is approached (plexus is not applicable 
because there are no anastomoses to be discovered between filaments; 
this is absolutely true for the coarse processes, but the terminal fila- 
ments from their very tenuity preclude the possibility of certainty). The Cerebellar Cortex of the Dog. 
209 
Many of the finest, like those in lower planes, turn downward in the 
process of division, but a large number approach the surface, still 
dividing, until they almost reach the sub-pial limit, where they 
apparently end. Doubtless this is the fact with many of them, but 
in sections taken at a somewhat oblique angle to the surface, a por- 
tion may be seen to turn at a different angle from which they arose, 
pass a minute distance nearly horizontally, and descend again, to be 
lost before reaching the middle third of the layer. Numerous darkly 
colored points (Flemming-hematoxylin, copper precipitate) at the 
ends of many of the filaments attest that a considerable portion 
through the whole layer change the angle of their course, their 
divided ends showing more deeply colored than the other part of the 
process, because it is now turned toward the objective and the 
observer is looking through a deeper bit of protoplasm—hence the 
intense stain. 
Variations and individual differences are of course not wanting 
in the manner of branching. One of the most common is for a cell 
to throw two or three thick primary branches upward past the middle 
third, but comparatively few filaments being given off at first by 
them; then past the centre, or even higher in the layer they divide 
interminably, or still proceeding upward to within 40 to 50(u of the 
surface, they throw off a fine spray of rarefied filaments, that, after 
approaching nearer to the “limitans externa” at an oblique angle, 
again pass off, descend, and are lost. Another peculiarity of the 
Purkinje cells which has been previously pointed out is that they 
spread out only in one direction. If the section is made in the long 
diameter of the convolution, yet perpendicular to the surface, it will 
be seen that the branches extend but little beyond the thickness of 
the cell itself; or, differently expressed, the rows are set like well 
trimmed osage-orange hedges planted closely together, and only 
capable of free expansion in a single direction—upward, and within 
the free space between their bodies,—though like the trees of the 
hedge there is much interlocking and crossing of the branches. 
Beyond the great cell layer, extending as high as the limit of the 
inferior third of the molecular layer, lie considerable numbers of 
small, rounded and angular multipolar cells, with distinct protoplasm, 
large nuclei containing coarse grains, and well developed prolonga- 
tions. The largest of them are 18 Xl2// and are undoubtedly nerve 210 
Henry J. Berkley. 
cells. In their neighborhood, but peripherally, are numbers of still 
smaller cells with a relatively large nucleus, and ill-defined though 
present protoplasm. These do not belong to any of the glia types, 
and are almost certainly, like their larger neighbors, nerve cells. 
It is in the immediate vicinity of both of these cell varieties that 
the nerve fibres that cross the “ limitans interna ” pass and disap- 
pear ; no connection is to be ascertained between them, yet from 
their being perfectly visible until they reach the neighborhood of the 
nerve bodies, then rapidly attenuate and are lost, a connection between 
the two is more than probable. A few fibres continue on beyond the 
inferior third of the zone, but they soon become non-medullated and 
disappear; but four medullated fibres in some 50 Weigert sections 
continued as far as the sub-pial periphery or to its vicinity. Some 
non-medullated fibres in inconsiderable numbers may occasionally 
be seen lying along the outer border of the band of tangential fibres, 
among the Purkinje cell extensions (safranin). 
The nuclei of the neuroglia in the barren layer are distributed 
throughout its central and mid-regions in rather less number than in 
the white tissue. Toward the pia, nuclei become less and less 
obvious, especially upon the vertex of the leaflet, where in places for 
comparatively large areas they are entirely absent; the entire space 
being filled, almost exclusively, by the ultimate ramifications of the 
Purkinje cells, capillaries, and supporting trabeculae of the pial con- 
nective tissue framework. The oval form predominates among these 
nuclei. Very many of their poles, with a little granulated proto- 
plasm attached, are directed vertically, and send out fine fibrilke 
upward and downward, crossing others that come from less frequent 
cells lying horizontally. Some of these glia fibrils are sufficiently 
coarse to allow of being traced to the external limiting membrane, 
and to the region of the pyramidal nuclei around the great cell row. 
All of these fibres, together with the capillaries, help to fill up the 
small remaining space between the Purkinje cell prolongations, and 
support the soft substance. 
The “Limitans Externa” and Piae Connections. 
On the outermost border of the leaflets subjacent to the pia is a 
condensation of substance marked by a deeper staining with certain 
reagents, notably nigrosine-carmine. With absolute alcohol (Nissl) The Cerebellar Cortex of the Dog. 
211 
and Flemming sections, this condensation is seen to be owing to a 
feltwork of fine glia fibrils, which are derived from a number of 
oval and pyramidal nuclei here situated. By no other process of 
hardening and staining can these nuclei be made sufficiently con- 
spicuous to be clearly seen. They vary greatly in number, the row 
in some spots being quite thickly set, in others very widely scat- 
tered (Fig. 7). From a part of these nuclei the feltwork is derived, 
while others contribute many fibrillse that descend, some to be lost 
in the higher portions of the layer, some to descend as far as the 
pear-nuclei row and terminate among them in the glia network. 
Between them can be seen, occasionally, triangularly-shaped bodies, 
from which descend stouter threads than from the pyramidal nuclei, 
which are continued directly, waving through the interposed elements, 
down to the pear-shaped and round cells of the Purkinje cell row, 
from which they apparently arise, and have their termination in 
these triangular bases, which are only a thickening of the glia sub- 
stance. 
Outwardly from this limiting membrane lies a sub-pial lymph 
space of considerable magnitude, crossed by rather thick connective 
tissue threads, that arise without much thickening at the point of 
origin, from the innermost layer of the vascular pia, crossing the 
lymph space, pass directly through the outer limiting membrane, 
and then through the molecular layer to the glia cell layer at its 
base, among whose fibres they also terminate. In the substance of 
the layer they are only to be distinguished by their larger calibre 
from the fibres arising from the glia cells (Fig. 7). 
The arrangement of the outer limiting membrane and the pial 
trabeculae is best seen between the lobules where the component 
parts are more spread out than on the surface; especially is this true 
of the trabeculae which in the depths of the furrows arise in such 
quantities as to form a brush-like figure, giving the most efficient 
support to the fragile nerve tissues of the cerebellar cortex. 
The number of nerve fibres entering into or passing from the 
molecular layer precludes the possibility of the termination of every 
large branch of a Purkinje cell becoming ultimately a nerve fibre, 
as has been supposed, and in fact there is no necessity to believe 
that more than one medullated nerve fibre can arise from a single 212 
Henry J. Berkley. 
cell, either physiological or histological, every evidence being to the 
contrary ; and all the centripetal and centrifugal coursing fibres must 
have their beginning in a single cell, their ending admitting of some 
doubt. However, there are sufficient nerve cells in the molecular 
and Purkinje cell layers to accommodate quite the number of fibres 
that are to be seen. 
The nerve elements of the cerebellum viewed as a unit bespeak a 
sensory and not a motor organ. The great cells of Purkinje have a 
closer likeness to those of Clarke’s columns, which are supposed to 
preside over a department of sensation, than to any other bodies in 
the nervous system. The round granules correspond in some 
measure to the granule layers of the retina, and the small cells of the 
molecular layer are like those in the posterior horns of the cord; all 
correspond with conceived ideas of sensory cells, and only a few 
multipolar bodies have any approach in appearance to motor cells. 
Lesions of the cerebellum, with the exception of the middle lobe, 
are without objective symptoms. It is conceivable that an involve- 
ment of the cortex of the middle lobe may also be symptomless, and 
that the true reason of the perversion of the sense of equilibrium in 
disease of this part is because the multiform crossings of fibres to 
and fro from one part of the organ to another are intersected, and 
that the symptoms arise, not from lesion of the cortex of the vermis 
or middle lobe, but from the destruction of the function of these 
crossing fibres, which then produces a loss of function, not of the 
middle lobe solely, but of the whole cerebellum. 
In the whole series of sections from all parts of the organ no 
essential difference was to be discovered between any of the different 
regions. 
March, 1891. 
Explanation of the Plate. 
Fig. 1. Skeleton drawing of one-half a cerebellar leaflet, built up 
from safranin, carmine, Weigert’s hematoxylin, Flemming-Weigert 
hematoxylin, carbolic fuchsin, Nissl’s magenta, and sodium-carminate 
sections, a, central core of white medullated fibres; al, glia nuclei 
among them; b, layer of granule or hematoxylin cells (all the eosin 
cells are not represented in this portion of the drawing); c, the 
granule layer with the eosin cells, straight nerve fibres running to The Cerebellar Cortex of the Dog. 
213 
the Purkinje cells, and the anastomosing fibre plexus; d, d, eosin 
cells; e, the fibre plexus; /,/, the straight fibres running through 
the granule layer, those passing directly into the molecular layer are 
not represented ; g, tangential fibres at the outer edge of the granule 
layer; h,li, the Purkinje bodies with their outspreading branches; 
hi, A2, turning of the terminal filaments beneath the pia; i, the pia 
covering the surface of the leaflet; k, nuclei in the sub-pial space; 
l, l, the small nerve cells in the inferior third of the molecular layer; 
m, fibres radiating out of the granule layer; n, nerve cells in the 
granule layer; o, the row of pear and round nuclei at the outer edge 
of the granule layer; ol, the Purkinje cell space, capsule not drawn ; 
p,p, glia elements of the molecular layer; o, o, fibrillse passing off 
from the pear-shaped cells attached to the limitans externa; r, sup- 
posed glia nucleus in the granule layer ; s, the limitans externa ; 
t, granule nuclei. Zeiss apochromatic immersion 1-12, ocular 4, 
reduced. 
Fig. 2. The grouping of the granule cells, Nissl’s magenta, a, a, 
granule cells ; b, larger clear nucleus belonging to the glia elements. 
The clear spaces between the granules are filled in by eosin cells, 
glia cells and medullated fibres. 
Fig. 3. A Purkinje cell divided through its centre to show the 
relation of the capsule and the cell body. Capsule a, b, from copper 
precipitate, nigrosine and safranin; medullated fibres and axis 
cylinder from Weigert, Flemming-Weigert, and safranin ; cell body 
from carmine and magenta; striations from nigrosine; nucleus and 
nucleolus from magenta. c, glia nuclei, the threads from whose 
bodies form the capsule, from magenta, safranin, and carbol-fuchsin 
preparations, c, c, medullated fibres. 
Fig. 4. Eosin cells, granule layer with the medullated nerve 
fibres, a, a, coursing over and by them, anastomosing and branch- 
ing at different points; b, coarser medullated fibre; c, large granules 
in the body of the eosin cell. Copper precipitate preparations. 
Fig. 5. 1, 2, 3, 4, 5, small multipolar and bipolar cells from 
the inner third of the molecular layer; 6, nerve cell with large 
nucleus and ill-defined protoplasm and extensions; from the granule 
layer: 1, carmine-nigrosine ; 2, 3, aqueous fuchsin; 5, 6, magenta. 
Fig. 6. Nuclei from the granule layer, showing the arrangement 
of the chromatin particles. 1, 2, 3, 4, neuroglia nuclei; 5, 6, nuclei 214 
Henry J. Berkley. 
of the hematoxylin cells, showing a minute ring of protoplasm around 
the nuclear substance. Magenta, nigrosine. 
Fig. 7. The cerebellar pia with its connections. a, the inner 
layer of the pia, a few nuclei are seen in the meshes; 6, the lymph 
space between the pia and limitans externa, with the meshwork of 
fibres and oval nuclei; c, the “limitans externa,” made up of fine 
threads arising from the oblong and round nuclei; d, e, stout fibres 
arising from the pia, descending entirely through the breadth of the 
molecular layer. Magenta and carbol-fuchsin sections. All the 
drawings were made with the aid of a Zeiss microscope, ocular 4, 
apochromatic immersion 1-12. THE JOHNS HOPKINS" HOSPITAL REPORTS. 
VOL. Ill PLATE III. 
F,g. I. 
Fig.2 
RS.3 
F.g-^ 
Tig 5. 
Fig 6 
Fi# 7 
CE i IE BE I, LAE C GET E X. 
A.Ho«*n A Co. Lith. Dailimnri*.