GRAFTING AND REGENERATION
By uniting parts of the same or different animals, or of plants, there
is given an opportunity of studying a number of important problems
connected with the regeneration of the grafted parts. Trembley’s
experiments in grafting pieces of hydra are amongst the earliest
recorded cases of uniting portions of different animals, although in
plants the process of grafting has been long known.[77] Trembley found
that if a hydra is cut in two, the pieces can be reunited by their
cut-surfaces, and a complete animal results. No regeneration takes place
where the union has been made. He also succeeded in uniting the anterior
half of one individual with the posterior half of another individual,
and again produced a single individual. He failed to obtain a permanent
union between different species.
More recently, Wetzel has carried out a number of different experiments
in uniting pieces of hydra. He found that if two hydras are cut in two,
the two anterior pieces may be united by the aboral cut-surfaces (Fig.
46, _B_), and the two posterior pieces may also be united by their oral
cut-surfaces (Fig. 46, _A_). The fusion of these “like-ends” takes place
as readily as when unlike ends are brought in contact, as in Trembley’s
experiments. Subsequently, however, regenerative changes take place.
When, for instance, two anterior pieces are united by their aboral ends,
there develop after two or three days one or two outgrowths, at or near
the line of union, that become new feet, and the two individuals may
subsequently separate. When two posterior pieces are united by their
oral surfaces, a double circle of tentacles generally develops, one on
each side of the line of union. The pieces then pinch apart and produce
two hydras.[78] In another experiment the head and a part of the foot
were cut from a hydra, and the head was turned around and grafted by its
aboral surface upon the aboral surface of the middle piece. Another
animal was cut in two in the middle, and the posterior half was grafted
by its oral end to the oral end of the middle piece. In this way a new,
artificial individual was made, as shown in Fig. 46, _C_, with the
middle part of the body in a reverse direction as compared with the
orientation of the two end-pieces.[79] The union of the three pieces was
so perfect that not even a swelling or a constriction indicated the
places of fusion. After six days a normal bud appeared at the region of
union of the posterior and middle pieces, that gave rise to a new hydra,
which separated after a few days. The compound animal was healthy and
ate many daphnias. It was kept under observation for twenty-four days,
and appeared normal, giving off several more buds.
[Illustration: FIG. 46.--_A._ Two posterior pieces of hydra united by
their oral ends. _B._ Two anterior pieces of hydra united by their
aboral ends. _C._ A “long hydra” made by uniting three pieces; the
middle piece reversed. _D._ After Peebles. Two posterior pieces of brown
hydra united by oral ends, and one cut off near union. A new anterior
end developed from the cut, aboral surface. _F._ After Peebles. Union of
a nutritive and a protective polyps of hydractinia. Subsequently former
cut off at line, 1-1. _E._ Union of two posterior pieces of hydra by
oral ends. Subsequently one piece cut off at line, 2-2. _E¹._ New head
regenerated in region of union, and a foot from aboral cut-end. _E²,
E³._ Fusion of two parts with a single hydra.]
In other experiments of this same sort a foot generally developed where
the two aboral surfaces came together, and the head-end separated from
the rest of the piece. In another case a mouth and tentacles appeared at
the place at which the oral ends had united.
In a different kind of experiment, the anterior ends of two hydras were
cut off and united by their aboral surfaces; then one of the components
was cut in two, just back of the circle of tentacles. After five days
two short, hook-like processes appeared at the cut, oral end. They
produced a foot, by means of which the animal fixed itself. In this case
it will be seen that a foot developed from an oral end. The result might
not in itself be considered sufficient to show whether the development
of a foot at the oral end of a piece is due to the influence of the
other component, or is simply a case of heteromorphosis having no
connection with the presence of the other component. Since
heteromorphosis has never been observed in isolated pieces of hydra, the
probability is that the result is in some way connected with the
presence of the other component. Peebles has made a number of
experiments, in which special attention was paid to this point. Fifteen
anterior pieces were united in pairs by their aboral cut-surfaces, and
then one component was cut in half, leaving an exposed oral end. Out of
this number five pieces formed a new head at the cut-surface, and the
pieces became attached by a foot, that developed at the region of union.
Two others did not regenerate at the cut-surface, but became fixed as
before, and neither regenerated nor became fixed at the cut-end. Three
became attached at the cut, oral surface, but none of these developed a
characteristic foot. The result shows, nevertheless, that some influence
was present that inhibited the development of a mouth and tentacles at
the oral cut-end, since these always develop in isolated pieces. In
another series of experiments posterior ends were united by their oral
surfaces, and then one of the two pieces was cut in two (Fig. 46, _E_).
A new hypostome and tentacles developed at the region of union, and a
foot at the aboral cut-surface, as shown in Fig. 46, _E¹_. An organism,
with one mouth and a circle of tentacles, and two bodies and two feet,
resulted. The bodies soon began to fuse together (Fig. 46, _E²_) into a
single one, and when the fusion had extended to the region of the feet,
they also fused into a single structure (Fig. 46, _E³_), so that a
single hydra was produced.
In another experiment, twenty-two posterior ends were united in the same
way, and then one of the two components was cut in two. In five cases a
single head developed on the aboral end of the smaller piece (Fig. 46,
_D_). It is evident in this case that the union of the two pieces has
been a factor in bringing about the development of an aboral head.
Another of the grafts produced an aboral head, and also one in the
region of union. The remaining sixteen grafts produced new heads, if
they developed at all, only in the region of union. Peebles states that
the regeneration of aboral heads takes place only when one component is
cut off near the region of union of the two pieces.
In general, it may be stated in regard to these experiments in hydra
that when pieces are united in the same direction, that is, by unlike
surfaces, a single individual is formed and no regeneration takes place
where the union has been made, but when like surfaces are brought
together, although perfect union may result, a process of regeneration
takes place later, at or near the line of union. Even the presence of
cut-surfaces at one or both cut-ends of the united components does not
generally affect the result, although, in a few cases, it may change it,
in so far that heteromorphic regeneration may take place from one piece.
This sometimes leads to a suppression of regeneration at the line of
union. The experiments do not show, perhaps, conclusively whether the
heteromorphosis of the smaller component is due to the polarity of the
larger component effecting a change in the smaller one, or whether the
closing of the oral end of the smaller component (by its union with the
other) brings about the result. All things considered, it seems to me
that the larger component has directly influenced the other.
King has found that if two posterior pieces of hydra are united by the
oral cut-surfaces, and then after they have fused _both pieces_ are cut
off _near_ the line of fusion, there develops from the small piece a
_single hydra_, with a foot at one end and tentacles at the other. If
only one of the pieces is cut off _near_ the line of fusion, a new head
develops from its oral surface, as Peebles had found. If two anterior
ends are united by their aboral cut-surfaces, and then later both are
cut off near the line of fusion, a single hydra develops from the small,
double piece. If one of the components is cut off near the line of
union, a foot develops from the oral cut-end. If in any of the cases the
cut is made some distance from the line of union, then each cut-surface
develops its typical structure. These experiments leave no doubt as to
the influence of the larger piece on the smaller one. Especially
interesting is the formation of one individual from two short pieces
united in opposite directions. In this case we must suppose that one
piece has the stronger influence on the combination (perhaps because it
is a little larger), and determines the polarization of the other piece.
King finds that when two posterior pieces are united by their oral ends,
regeneration of one or of two heads often takes place at the line of
union (Fig. 47, _B, B¹, B²_), as Wetzel had found. If a dark green
individual is united to a light green one, it can be seen that in many
cases the new heads are formed by both components, as shown in Fig. 47,
_B_¹. Later one of the posterior ends is absorbed, and the halves may
then separate (Fig. 47, _B¹, B²_). If a number of pieces are united,
as indicated in Fig. 47, _E_, a number of heads may be formed, and one
or more of these may have a double origin. No evidences of separation
of the pieces was observed in cases of this sort.
[Illustration: FIG. 47.--After King. _A._ Hydra split in two, hanging
vertically downwards. Later the halves completely separated. _B._ Two
posterior ends united by oral surfaces. _B¹._ Same; it regenerated two
heads, each composed of parts of both pieces. _B²._ Absorption of one
piece leading to a later separation of halves. _C._ Two posterior ends
united by oblique surfaces. Later one piece partially cut off, as
indicated by line. _C¹._ Later still, two heads developed, one at _N_,
the other at _M_. _D._ Similar experiment in which only one head
developed, at _M_. _E._ Five pieces united as shown by arrows. Four
heads regenerated, one being composed of parts of two pieces.]
In one experiment two posterior pieces were united by oblique surfaces,
as shown in Fig. 47, _C_, and one of the two was afterwards cut across,
as indicated by the cross-line. The subsequent regeneration that took
place is shown in Fig. 47, _C¹._ A head, composed of parts of both
pieces, developed at the cut-surface _M_, and another in the region _N_
in Fig. 47, _C_, composed of material of one component. In another case,
shown in Fig. 47, _D_, a head developed only at the cut-edge, but it was
made up of material from both components.
A series of grafting experiments of another sort has been made
[Illustration: FIG. 48.--After Rand. _A._ Head of Hydra cut off. After
eight days. _A¹._ Same after thirteen days. Three tentacles misplaced.
_A²._ Same after eighteen days. _A³._ Same after twenty-one days.
Misplaced tentacles absorbed. _B._ Anterior end of _Hydra fusca_,
grafted upon side of body of another individual. Half an hour after
operation. _B¹._ Same after four days. _B²._ Same after thirty-eight
days. _B³._ Same, foot-region after forty-nine days. _B⁴._ Same after
separating. Fifty-second day.]
by Rand. A part of one hydra is grafted upon the side of another one in
the following way. A groove is scratched in a film of soft paraffine
covering the bottom of a dish filled with water. Another groove is made
at right angles to the first one, and opening into it. A hydra (the
stock) is placed in the first groove, and a wound made in its side with
a knife. Another hydra is cut in two, and one piece (the graft) placed
in the other groove, and its cut-surface brought into contact with the
wound in the side of the first individual. If the operation is
successful the exposed surfaces of the two hydras quickly unite, and the
combination may be taken out of the groove. If the piece grafted on the
stock included about the anterior half of a hydra, a two-headed animal
results, as shown in Fig. 48, _B_. Although the graft has been united to
the side of the stock, it soon assumes an apparently terminal position
(Fig. 48, _B¹_). This is due to the graft sharing with the anterior end
of the stock the common basal portion of the stock. A slow process of
separation of the two anterior ends now begins, brought about by a
deepening of the angle between the halves (Fig. 48, _B²_). This leads
ultimately to a complete separation of the two individuals (Fig. 48,
_B³_, _B⁴_). Each may get a part of the original foot, or a new foot
may arise on the graft as the division approaches the base.
In other experiments only a small part of the foot-end was cut from the
animal that served as the graft. The long anterior piece was grafted as
before upon the side of the stock. After the two had united, the graft
was cut in two, leaving a part of the graft attached to the stock. The
part regenerated tentacles, and in two cases subsequently separated from
the stock as in the first experiment. In a third case the graft was
absorbed by the stock as far as the circle of new tentacles, but its
subsequent fate was not determined. In a fourth case the graft did not
regenerate its tentacles, and was completely absorbed into the wall of
the stock. The smaller the piece that is grafted on the stock the
greater the chance that it will be absorbed, and furthermore short,
broad rings are more likely to be absorbed than long, tubular pieces of
the same volume.[80]
Rand’s results show in general that when hydras are grafted together
they regain the typical form in one of two ways,--either by separation
into two individuals, or by the absorption of the smaller into the
larger component. In the former case the result is brought about in the
same way as when the anterior end is partially split in two and the
halves subsequently separate. When the graft is absorbed it is not clear
whether the absorbed piece disappears or, as seems not improbable, forms
a part of the wall of the stock.
It is important to notice the difference between lateral buds and
lateral grafts. The buds separate in the course of four or five days by
constricting at the base, but this never happens in lateral grafts. Rand
has also made some experiments with buds. He cut off the outer oral end
of a bud, and grafted it back upon the stock in a new place. It did not
separate from the stock as does a bud, but by a slow process of division
it was set free in the same way as are lateral grafts. The proximal end
of the bud, which was left attached, developed tentacles at its free
end, constricted at its base, and was set free. The separation was,
however, somewhat delayed.
In another experiment a bud was split in two lengthwise, and the cut was
extended so that the body of the parent was separated into two pieces.
Twenty-four hours later it was found that each half-bud had closed in,
and was much larger than when first cut. The half-bud, that was attached
to the posterior end of the anterior piece, was constricting at its
base, and subsequently it separated at its point of attachment. The
other half of the bud, that had been left attached to the anterior end
of the posterior piece, had swung around, so that its long axis
corresponded to that of the posterior, parental piece. At first a slight
constriction indicated the line of union of the two, but later this
disappeared and a single hydra resulted. Whether the difference in the
fate of the two half-buds is connected with their different polar
relations to the parts of the parent, or is due to some other difference
in the absorbing power of the anterior and posterior pieces, is not
known.
[Illustration: FIG. 49.--After Peebles. _A._ Grafting in _Tubularia
mesenbryanthemum_. A small piece of the stock taken from the region near
the base, and grafted in a reversed direction on the oral end of a long
piece. _B._ Same with distal tentacles in small piece, and proximal
tentacles in large piece (modified from Peebles). _C._ Same. Formation
of hydranth (original). _D._ Like _A_. Both pieces produce hydranths.
_E._ Protrusion of hydranths of last. _F._ Piece of oral end cut off,
turned around and grafted on oral end of long piece. A single hydranth
produced. Distal tentacle from both components. _G_. A short piece from
distal (oral) end of long piece cut off, and grafted by its proximal end
to proximal end of the same long piece.]
Tubularia is not so well suited as hydra to show the influence of
grafting on the united parts, since pieces of tubularia produce
hydranths, both at the oral and aboral ends, although the latter
hydranths take longer to develop. Peebles has shown, nevertheless, that
grafting has an influence on the behavior of a piece. In order to show
that the polarity of a small piece could be affected by a larger piece,
the following experiment was carried out. After cutting off the old
hydranth from the end of a stem, a short piece was then cut from the
distal end of the same stem, turned around, and its oral end brought in
contact with the oral end of the original piece, as indicated in Fig.
49, _F_. The two pieces, being held together for a few minutes, stuck
together and subsequently united perfectly. From eighty-eight pieces
united in this way the following results were obtained. Thirty-six
formed a single hydranth at the end at which the grafting had been made.
The distal row of tentacles appeared in the smaller reversed component,
the proximal row in the larger piece (Fig. 49, _B_). The new hydranth
pushed out later through the perisarc of the smaller piece (Fig. 49,
_C_). In this experiment the smaller component was shorter than the
average length of the hydranth-forming region. In two cases, in which
the smaller component was larger, both circles of tentacles appeared in
this piece. In six of the experiments the tips of the proximal tentacles
arose from a part of the wall of the smaller piece, hence these
tentacles had a double origin (Fig. 49, _F_). In five of the unions the
smaller as well as the larger component produced a hydranth; the two
were stuck together by their oral ends (Fig. 49, _D_, _E_). The
remaining four unions gave somewhat different results. In three of these
the smaller piece produced only a part of a hydranth that remained
sticking to the end of the hydranth formed by the larger component. In
the thirty-six cases in which the minor component took part in the
formation of the single hydranth, the influence of the larger component
was shown not only in reversing the polarity of the smaller component,
although this might in part be accounted for by the closing of the oral
end of the smaller piece, but also in the time of development, since the
hydranth appeared sooner than does the aboral hydranth and at the same
time as does the oral hydranth.
In another series of experiments, a short piece was cut from the basal
end of a long piece (three to four centimetres) and brought forward and
grafted in a reversed position on the anterior end of the same long
piece (Fig. 49, _A_). Of five unions of this sort, one produced a
hydranth in each component, neither being reversed. Another of the
pieces produced a hydranth partly out of each component (and at the same
time another at the aboral end of the large piece). The other two pieces
produced a single hydranth, a part of which came from the minor
component and appeared before the aboral hydranth on the aboral end of
the larger piece. This last result shows that the small piece from the
basal end has been affected by the oral end in such a way that it
develops more rapidly than it would have done had it remained a part of
the basal end.
In a third series of experiments a short piece (about a half of a
millimetre) was cut from the anterior end of a long piece (one and
five-tenths to two centimetres) and grafted _in a reversed position_ on
the posterior end of the same long piece (Fig. 49, _G_). In four cases a
hydranth developed only at the oral end of the long piece and none from
the aboral end or from the short piece. Eight unions produced, however,
in the region of the graft, a hydranth formed partly by each component.
Later another hydranth developed at the oral end of the larger piece.
The latter results are not convincing, but they may show that the small
piece has hastened the development of the hydranth at the aboral end.
Peebles has also made some experiments in grafting pieces of different
members of the colonies of hydractinia and podocoryne. The colony of the
former is made up of three different kinds of individuals: the
nutritive, the reproductive, and the protective hydroids. A series of
preliminary experiments showed that if these individuals are cut into a
number of pieces each piece regenerates the same kind of individual as
that of which it had been a part. It was also observed that if pieces of
the nutritive individuals were allowed to remain quietly on the bottom
of the dish they sent out branching stolons, which stuck to the bottom
of the dish, and from these stolons there arose later nutritive
hydranths that stood at right angles to the surface. When pieces of the
same kind of individuals are grafted together, the results are
essentially the same as with tubularia. If pieces of different kinds of
individuals are united, the opportunity is given of testing the possible
influence of one kind on the other. Peebles united a nutritive and a
protective polyp by the cut, aboral ends (Fig. 46, _E_), and after they
had grown together one of the polyps was cut off near the region of
union, so that a small piece of a nutritive polyp was left attached to a
protective polyp. When the piece of the nutritive polyp regenerated, it
made a new nutritive polyp. The influence of the protective polyp was
not apparent. If a nutritive and a reproductive polyp are united in the
same way, and the latter cut in two near the line of union, a new
reproductive polyp develops from the piece left attached to the
nutritive polyp. Again there is shown no influence of the one on the
other kind of polyp.
Hargitt has also made a number of grafting experiments on other
hydroids. His most interesting results are those in which parts of two
medusæ were united by holding their cut-surfaces together by means of
bristles passing through the individuals. Hargitt also finds that while
in certain hydroids it is possible to bring about a union of oral with
oral end, or aboral with aboral, or oral with aboral end of the same
species,[81] yet a permanent union between different species cannot be
brought about. These results are in agreement with those of a number of
writers who have recorded the difficulty or impossibility of uniting
parts of different species of hydra. In a few instances it has been
possible to unite temporarily a piece of a brown hydra with a piece of a
green one,--as I have also seen accomplished,--yet the pieces
subsequently separate. Wetzel succeeded in obtaining better results with
two species of brown hydras, _Hydra fusca_ and _Hydra grisea_. In one
experiment the head of _Hydra grisea_ was grafted on the body (from
which the head had been cut off) of _Hydra fusca_. After five hours the
pieces seemed to have united. Later a constriction appeared at the place
of union, and the head-piece produced a foot near the line of union, and
the posterior piece produced a circle of tentacles at its anterior end.
Eight days later, when the animal was being killed, it fell apart into
two pieces. It was observed that during the period of union a stimulus
to one piece was not carried over to the other. Wetzel’s results seem to
show that pieces of these two species of hydra unite at first, when
brought together, as perfectly as do pieces of the same species, but the
union never becomes permanent, a constriction appearing later at the
line of union, and the pieces separating in this region. These results
indicate, it seems to me, that the factors that bring about the first
union are different from those that make the grafted pieces one organic
whole. Other results indicate that the union of oral to oral end, or
aboral to aboral end, while at first as perfect as between unlike
surfaces, nevertheless is less permanent than when unlike surfaces are
united; at least, subsequent regeneration is more likely to occur in the
former than in the latter, and after this occurs the separation of the
individuals often takes place. It seems, moreover, not improbable that a
more permanent union results when similar regions are united by unlike
surfaces, than when the union is at different levels. If, for instance,
the anterior half of one hydra is united to the posterior half of
another individual, the union is generally permanent; but if one or both
of the pieces are longer than half the length, so that a “long animal”
results, new tentacles are more often formed at the oral end of one
component, and the parts subsequently separate. It may be that, at
present, the data are insufficient to establish this general rule, and
no doubt other modifying influences must be also taken into account; but
it is important that attention should be drawn to this side of the
subject.
Grafting experiments in planarians have so far been carried out in only
the two cases which I have described. In one of these the anterior ends
of two short pieces of _Bipalium kewense_ were united (Fig. 50, _A_).
Neither piece produced a head at the region of union. Later the pieces
were cut apart by an oblique cut that passed across the line of union
(Fig. 50, _C_), so that each piece retained at its most anterior end (at
one side) a piece of the other individual in a reversed position. A head
developed at the anterior (and lateral) end of each piece, in such a way
that a part at least of the small reversed piece was contained in the
new head (Fig. 50, _D_). In the other case two pieces of bipalium were
united by their posterior cut-surfaces. Each piece produced a new head
at its free end, and the pieces greatly elongated, but remained sticking
together (Fig. 51).
[Illustration: FIG. 50.--_A._ Two pieces of _Bipalium kewense_ united by
anterior ends. _B, C._ Later stages of same. Line in _C_ indicates how
pieces were cut apart. _D._ Two worms produced by these pieces. All
drawn to scale.]
[Illustration: FIG. 51.--Two pieces of _Bipalium kewense_ united by
posterior ends. Each regenerated a head at anterior end.]
A large number of experiments have been made by Joest in grafting pieces
of earthworms. The cut-surfaces were held in contact by means of two or
three threads passing through the body wall of each piece and tied
across, so that the pieces were drawn together and held firmly in that
position. Joest found that pieces of the same or of different
individuals could be united in various ways, and the union become
permanent. If the anterior end of one worm is united to the posterior
end of the same, or of another worm, a perfect union is formed, and no
subsequent regeneration takes place (Fig. 52, _A_). Long worms can be
made by uniting two pieces, each more than half the length of a worm, or
by uniting three pieces, as shown in Fig. 52, _C_. Short worms can be
formed by cutting a middle piece from a worm, and uniting the anterior
and posterior pieces (Fig. 53, _D_). Joest found that when a short worm
is made in this way, so that no reproductive region is present, the new
worm does not produce new reproductive organs. It is conceivable that
new reproductive organs might have been produced either in the old
segments, or by the formation of a new reproductive region between the
two united pieces, but neither process takes place. In the long worms
two sets of reproductive organs, etc., are present. This sort of union
is, however, less permanent, as the worms often pull apart.
[Illustration: FIG. 52.--After Joest. _A._ Union of two pieces of
_Allolobophora terrestris_ in normal position. Twenty-two months after
operation. _B._ Union of two pieces _Lumbricus rubellus_. Pieces turned
180° with respect to each other. _C._ Union of three pieces of _A.
terrestris_ to make a “long worm.” _D._ Union of two worms (by anterior
ends) from each of which eight anterior segments had been removed. After
three months. Regenerating two new heads. _E._ A small piece of
_Lumbricus rubellus_ grafted upon _Allolobophora terrestris_. Former
regenerated an anterior end.]
Joest also united two posterior ends by their anterior surfaces. In many
cases no regeneration took place, and, in the absence of a head, the
combination is destined to die, although it may remain alive, without
food, for several months. When two very long pieces were united by
their anterior ends,--only eight segments being removed from each
worm,--although perfect union took place at first, later one or two new
heads generally developed at the region of union (Fig. 52, _D_). When
only one head developed it did not seem to belong to one of the
components rather than to the other, and originated in the new tissue
that appeared between the two pieces. These experiments, in which the
anterior surfaces of two pieces are united, show also that the new head
arises between the two pieces most often, if not exclusively, when the
union is in the anterior ends of the worms. This corresponds with what
is now known in regard to the development of new heads by isolated
pieces, since there is less tendency to produce a head the farther
posteriorly the cut has been made. At more posterior levels a tail and
not a head is often regenerated, as has been stated, on the anterior
cut-surface. This formation of a heteromorphic tail seems to have been
suppressed in the pieces united in this region, except in one case,[82]
in which it appears, from Joest’s account, that a tail probably
regenerated, although Joest speaks of it as a head.
It is more difficult to unite two anterior ends by their posterior
cut-surfaces, not because the surfaces refuse to unite, but because the
two pieces crawl away from each other and pull apart. In one case,
however, union of this sort was brought about.
In all the combinations that have been so far described, the dorsal and
ventral surfaces of both components were kept in the same direction, so
that the ventral nerve-cord of one piece came in contact and fused with
the nerve-cord in the other piece. Sometimes it may happen that the
components are not quite in the same position, and the end of one
nerve-cord may fail to abut against the other one. In such cases Joest
thinks that regeneration is more apt to take place in the region of
union, and he has carried out a series of experiments in which the
pieces were intentionally united, so that they are not in corresponding
positions. It is found that if one piece is turned so that the nervous
system lies 90 degrees, or even 180 degrees (Fig. 52, _B_), from that of
the other piece, the union takes place just as when the pieces have the
same orientation, except that the ends of the nerve-cords do not unite.
Subsequent regeneration from one or from both components generally takes
place in the region of union.
It is more difficult to unite pieces of different species of worms, yet
Joest has succeeded also in making combinations of this sort. One union
between the anterior end of _Lumbricus rubellus_ and the posterior end
of _Allolobophora terrestris_ was permanent, and the new worm reacted as
a single individual, and lived for eight months. Each piece retained its
specific characters, and showed no influence of the other component. By
means of a similar experiment we have a way of finding out if one
component can influence regeneration taking place from the other piece.
Although Joest made only a few observations of this sort, the results
show that no such influence is manifested.
[Illustration: FIG. 53.--After Joest. _A._ Small piece of _Allolobophora
terrestris_ from posterior end grafted upon anterior end of another
individual. Oral end free. Four weeks after grafting eight new segments
formed. _B_. Same fourteen days later. A new part of thirty-seven
segments had appeared at end of former eight segments. _C._ A piece of
the body wall of _Allolobophora terrestris_ grafted upon the cut-end
(anterior) of _Lumbricus rubellus_. Two months later, as shown in
figure, a head had grown on major component. _D._ Anterior and posterior
ends of _A. terrestris_ united to make a “short worm.” _E._ A piece of
body wall of _A. cyanea_ grafted on side of body of _Lumbricus
rubellus_. _F._ Piece of _L. rubellus_ grafted on side of body of
another individual to produce a double-tailed worm.]
By means of grafting it is possible to keep alive small pieces of a worm
that would otherwise perish. For instance, pieces of a worm containing
only three segments are not capable of independent existence, except for
a short time, and even pieces of from four to eight segments die in most
cases. It is not possible to unite small pieces of this size directly
upon larger pieces, since they will die, ordinarily, as a result of the
operation, but larger pieces can be united and then after union has been
effected, one of them may be cut off near the place of union. The same
result is sometimes brought about accidentally by the worms themselves
pulling apart and leaving a small piece of one component attached to the
other. Joest found that in several cases these small, attached pieces
regenerated. In one case, after two long pieces had pulled apart, a
small piece, left by one of the two, regenerated a single new segment
with a mouth at its end. In another case, after one of the components
had been cut off, leaving two segments attached, a new part of seven
segments regenerated.[83] Especially interesting is the case in which
two individuals (_A. terrestris_) had been united to form a long worm.
The anterior component extended to within two centimetres of the anus;
the posterior piece had had the first four segments removed. Three days
later the anterior piece was cut off three segments in front of the
region of union. About a month later a small part of eight segments had
regenerated from the cut-end (Fig. 53, _A_). Fifteen days later another
new part of thirty-seven segments developed at the end of the first new
part (Fig. 53, _B_). Joest speaks of the first eight segments as a head,
and the second simply as a regenerative product. There can be little
doubt, I think, that both parts represent a heteromorphic tail. The
region from which the regeneration took place would make this
interpretation highly probable, and Joest’s figures also indicate that
the structure is a tail. The result is very interesting, if my
interpretation is correct, as it shows that the major component did not
influence the kind of regeneration, although the surface of regeneration
was separated by only three tail-segments from the anterior end of the
major component.
In another experiment a long animal was made by uniting _Lumbricus
rubellus_ (whose posterior third had been cut off) and _Allolobophora
terrestris_ (whose first six segments had been cut off). Four days later
the two components had torn apart, but a small piece of the anterior
worm remained attached to the anterior end of the posterior component.
The small piece consisted of the dorsal part of two and a half segments
without any ventral part, so that the anterior end of the posterior
component was partially exposed. The small piece of lumbricus was much
lighter in color, and this difference made it easy to distinguish
between the two. In less than a month the small transplanted piece had
replaced its missing ventral part, so that the entire anterior surface
of the larger component was covered over. The small piece, in addition
to regenerating its ventral part of four segments, had also begun to
make new segments. After a month and a half six new segments were
present (Fig. 52, _E_), with a mouth at the anterior end.[84] Even after
ten months the color of the small piece was strikingly different from
that of the major component. The new head had the typical red-brown
color of _L. rubellus_, that forms a strong contrast to the grayish blue
color of _A. terrestris_.[85] The result shows that the color of the
regenerated part has not been influenced by that of the posterior
component, and this is all the more interesting, as Joest points out,
because the small piece that was left after the worms pulled apart was
too small to have lived independently for any length of time, and must
have derived all its nourishment from the larger piece.
In other experiments pieces of one species were cut from the side of the
body and grafted upon the cut-surface of the anterior end (or elsewhere)
of another species. In one of these experiments a piece from the side of
_A. terrestris_, that extended over five or six segments, was sewed upon
the anterior cut-surface of _L. rubellus_ (from which the anterior five
segments had been removed). In about a month new tissue appeared on the
ventral side between the two pieces, and a little later a complete head
developed, whose dorsal side was made up of the small piece (Fig. 53,
_C_). The grafted piece was dark, and the new, regenerated part light in
color and continuous with the brown color of _L. rubellus_, from which
the new part had arisen. It is important to notice that the four
segments of the graft are completed by four segments of the new part.
After three months the new part had assumed the red-brown color of _L.
rubellus_. The color of the grafted piece had not changed. We see in
this case that even the presence of a part of another worm in a
regenerating region does not have any influence, at least so far as
color is concerned, on the new part, even though its segments supplement
some of those of the new part. The new tissue seems to have come
entirely from the major component, and to have carried over the color
characteristics of the old part.
It has been shown that when two posterior pieces are united by their
anterior ends the combination must sooner or later die, since it has no
way of procuring food. The question arises: What will happen if one of
the two components is cut in two near the place of union? Will a head
then develop on the exposed aboral surface, _because_ a head is needed
to adapt the worm to its surroundings, or possibly, if it occurred,
because the major component exerts some sort of influence on the short,
attached piece, as happens in hydra and in tubularia? Both Joest and I
carried out an experiment of this sort, and found that a tail and not a
head regenerated, as shown in Fig. 16, _F_. The experiment is, however,
insufficient to answer the question, since the region in which the
second cut was made is a region from which only a tail (and not a head)
arises, even when the oral end of a piece is exposed. In order to avoid
this difficulty I carried out another experiment. Two worms had the
first five or six segments cut off and the exposed anterior ends of the
worms united, as shown in Fig. 16, _D_. Then one of the components was
cut off, leaving three or four segments attached to the anterior end of
the other component. Although regeneration began in one case, it did not
go far enough to show what sort of a structure had developed, but Hazen,
who took up the same experiment, succeeded in one case in obtaining a
definite result. At the exposed aboral end of the small piece a head and
not a tail developed (Fig. 16, _E_). At first sight it may appear that
the result shows the influence of the major component on the small
piece, causing it to produce a head and not a tail at its aboral end,
but I think that this conclusion would be erroneous, because it seems
much more probable that we have here a case of heteromorphosis, similar
to that in _Planaria lugubris_, and that the result depends entirely on
the action of the smaller component. It is hardly possible to
demonstrate that this is the correct interpretation, since if a small
piece of this size is isolated it dies before it regenerates. The result
is paralleled, however, by the regeneration of a tail at the anterior
surface of a posterior piece.
The process of grafting has long been practised with plants, but the
experiments were made more for practical purposes than to study the
theoretical problems involved. Vöchting has, however, carried out a
large number of well-planned experiments. He finds that a stem can be
grafted upon a root, and a root upon a stem, a leaf upon a stem or upon
a root. Even an entire plant can be grafted upon another. The results
show, however, in general, that, whatever the new position may be, the
graft retains its morphological characters--a shoot remains a shoot, a
root is always a root, and a leaf a leaf. Vöchting concludes that there
is in the plant no principle or organization that conditions an
unchangeable arrangement of the main organs. “The inherited order of the
parts, acquired apparently on physiological grounds, may be altered by
the experimentator; it is possible for him to change the position of the
building blocks within a wide range without endangering the life of the
whole.” “It is essential, however, for the success of the experiment
that the grafted parts, or tissues, retain their normal orientation. If
this condition is not fulfilled there may take place, it is true, a
union of the parts, but sooner or later disturbances set in.” Vöchting
transplanted pieces in abnormal positions, sometimes reversing the long
axis of the grafted piece, sometimes the radial axes, and sometimes both
together. In some cases this led to the formation of swellings that
interfered with the nourishment but carried with it no further
consequences. In other cases the changes went so far that the vital
processes were interfered with. At times an incomplete union took place
between the parts; at others, even though the first union was perfect,
death later ensued.
On the other hand, when similar pieces were grafted with their original
orientation, a perfect union took place and the piece became a part of
the stock. The results establish, Vöchting claims, that every part and
every portion of a part has a polar orientation in one direction, and
furthermore, in a body having a radially symmetrical form, there is also
a radial polarization; that is, the inner side of each part is different
from the outer side of the same surface, even though no such difference
is apparent to us. The properties of the tissue-complex rest, in the
last analysis, on that of the cells; the properties of the whole being
only the sum total of the properties of its elements, so that we may say
that every living cell of the root is polarized, not only
longitudinally, but also radially; each has a different apical and root
pole, a different anterior and posterior pole, and also right and left
polar relations. These results, deduced from the experiments in
grafting, lead Vöchting to formulate the following rule: “Like poles
repel, unlike poles attract.” This rule is the same as the law of the
magnet. In fact, Vöchting states that the root and the stem relations
show a remarkable resemblance, despite many differences, to a magnet. If
the magnet is broken into pieces it may be reunited by bringing unlike
poles together, but not by uniting like poles; the same statement holds
for the root and the stem.
Exception may be taken, I believe, to parts of Vöchting’s conclusions,
especially in the light of the recent experiments in grafting in
animals. It is by no means to be granted without further demonstration
that the properties of the whole organism are only the sum-total of the
action of the individual cells. If, as seems to be the case, the cells
are organically united into a whole, the properties of this whole may be
very different from the sum of the properties of the individual cells,
just as the properties of sugar are entirely different from the sum of
the properties of carbon, hydrogen, and oxygen.
The statement that like poles repel and unlike poles attract is, I
believe, a conclusion that goes beyond the evidence. The experiments
show that like poles do often unite in plants, and this has been
abundantly shown to be the case in the lower animals, and even in forms
as high as the earthworm and the tadpole. Even if when like poles are
united subsequent changes take place, that in some cases, although
apparently not in animals, lead to the death of the graft, it by no
means follows that this has anything to do with the attraction or
repulsion of the parts, but rather with some difficulty in obtaining
food, or with the transportation of substances through the plant. In the
lower animals we have seen that when like poles are united there is
sometimes a stronger tendency to produce new organs at or near the place
of union than when unlike poles are united, but it would be going too
far, I think, to state that this is due to repulsion of the parts,
especially in the sense in which the like poles of a magnet repel each
other. It seems to be due rather to the two parts failing to unite into
a whole organization, each retaining the same structural basis that it
had before grafting, but this is a very different principle from that of
an attraction and repulsion of the parts, and the question of the union
of the parts appears also to be a different question from that of the
organization of the parts themselves.
In the mammals, and in general in all forms in which there is a
dependence of the parts on each other, it is impossible to carry out
grafting-experiments on the same scale as those described in the
preceding pages. The principal difficulties are to make the parts unite,
and to supply nourishment and oxygen to the graft. Owing to the
dependence of the parts of the body on each other for a constant supply
of oxygen and food derived from the blood, as well as for the removal of
the waste products, the parts cannot remain alive, or even in good
condition, while new connections are being established. For this reason,
as well as for others, it would not be possible, for instance, to graft
the arm of a man upon another man. The tissue may have the power of
uniting even in this case, as is seen when the bone is broken and
subsequently reunited, but the difficulty would be in supplying the
grafted arm with nourishment, etc., during the long time required for
the union to take place. Smaller parts of the body may be successfully
grafted, and there are several recorded cases in which parts of a
finger, or of the nose, are said to have been cut off and to have
reunited after being quickly put back in place. Pieces of human skin may
be grafted without great difficulty upon an exposed surface, and it has
been said that small pieces succeed better than larger ones, owing, most
probably, to their being able to absorb sufficient oxygen, etc., and
keep alive until new blood vessels have grown into the grafted piece.
There are a number of old and curious observations in regard to cases of
grafting in higher animals. It was found by Hunter and by Duhamel that
the spur of a young cock could be grafted upon the comb, when it
continued to grow to its normal size. The comb, being richly supplied
with blood, furnished the nourishment for the growth of the spur.
Fischer transplanted the leg of an embryo bird to the comb of a cock, or
of a hen, where it grew at first, but after some months degenerated.
Zahn transplanted the fœtal femur to the kidney, where it grew for a
time, but later degenerated. Bert transplanted the tail of a white rat
to the body of _Mus decumanus_, where it continued alive; but he found
that the tail of the field mouse, _Mus sylvaticus_, did not grow so well
on the rat, and the tail of a rat would not unite at all with the body
of a dog or of a cat. Bert bent over the tip of the tail of a rat, and
grafted the distal end into the skin of the back of the same animal.
After the tip had established union with the surrounding tissues, the
tail was cut off at its base. The grafted tail remained alive, but did
not regenerate at its free end.
There are several cases described by pathologists in which the skin of
one mammal has been transplanted to another. The transplantation of the
skin of the negro upon a white man has been brought about, but the
evidence as to what subsequently happened is contradictory. It appears
that while in many instances the transplanted skin has remained alive
for a time, yet later it was thrown off by new skin growing under it and
replacing it.
Leo Loeb has described a curious instance of grafting pieces of skin of
different colors in the guinea pig. If a piece of black skin from the
ear of a guinea pig is grafted upon the white ear of another animal, it
unites and continues to live, but if a piece of white skin is grafted
upon a black ear, it is slowly thrown off and replaced by new black skin
that has regenerated around the edge of the graft from the tissue of the
black ear.
In the literature of pathology there are many cases described in which
parts of the body of mammals, particularly internal organs, have been
grafted in unusual regions. The results have not always been the same,
for while in some cases it appears that the operation has succeeded, in
others the grafted part is subsequently absorbed, and in still other
cases the graft may be at first partly absorbed and later begin to grow
again. It appears that the establishment of an adequate blood supply is
the most important element of success. Ribbert, who has made an
extensive and successful series of experiments, has stated that the
grafting takes place better when small pieces of an organ are used,
since these can draw immediately on the surrounding regions for their
oxygen, etc., while larger pieces are found to break down in the
interior, owing to the fact that this part is too far removed from the
supply of oxygen, food, etc. After the grafted piece has established a
blood supply of its own, it may continue to grow. Ribbert transplanted
small pieces of different tissues of the rabbit and guinea pig in, and
upon the surface of, the lymph glands of the same or of another
individual. The lymph gland was chosen because small pieces of tissue
can be afterwards easily detected. A small piece of tissue about as
large as a pin’s head is cut off from whatever tissue is to be grafted,
and as quickly as possible placed in a small cleft made in the lymph
gland. After several days, weeks, or months, the gland is removed and
the graft examined by means of serial sections.
Most of the experiments were made with “epithelial organs,” and
according to Ribbert, if pieces of such organs are composed of
epithelium only, they cannot be successfully grafted. For instance, the
cells of the cornea can be readily separated from their underlying
connective tissue, and can be kept alive in the lymph gland, but the
cells diminish in number, show retrogressive metamorphosis in the
direction of atrophy, and are finally absorbed. It seems that epithelium
by itself cannot extract nourishment from its surroundings. Nothing is
easier, however, than to transplant epithelium, if its connective tissue
is present. The connective tissue furnishes so good a basis for
nourishment that the epithelium not only lives, but may continue to
proliferate. Ribbert finds that pieces of skin roll in after their
removal. Then a process of growth takes place corresponding to that
which follows a wound in the skin. The surface is closed and a small
cyst is formed with a central cavity. The epithelium undergoes no
changes during the first days or weeks. It remains stratified and shows
an active process of cornification and desquamation. Similar results
were obtained when pieces of the conjunctiva were transplanted, either
under the skin in the anterior chamber of the eye, or in the lymph
gland.
A small piece of the lining epithelium of the trachea with its
underlying cartilage was also placed in the lymph gland. The epithelium
grew, and covered over the wounded surface, forming over it only a
single layer of cells. The old many-layered epithelium also became
arranged in a single layer.
The wax glands, found in the inguinal folds of the rabbit, were also
transplanted. The gland is composed of closed, compressed alveoli,
surrounded by large, polygonal, clear cells. Small pieces of a gland,
transplanted upon the lymph gland, underwent characteristic changes. The
cells of the alveoli were changed into a stratified epithelium; and
broken-down cells, and wax, were found in the interior of the alveoli.
The central alveoli underwent the greatest change, while some of the
peripheral alveoli that were in contact with the lymph gland remained
unchanged. It seems that the difference is due to the better nourishment
of the outer alveoli. After several months the alveoli swell up and
degenerate. Transplanted pieces of the salivary glands also change, the
alveoli producing a lining epithelium like that of the transplanted wax
gland. The same change was observed in a piece of a salivary gland
transplanted in the body cavity.
Small pieces of the liver were cut off and placed in the lymph gland.
They did not always grow as well as did the preceding tissues, but often
went to pieces. If they healed, the liver tissue often remained
unchanged for several weeks. After two or three weeks connective tissue
appeared between the peripheral liver cells, separating the cells from
each other. The cells grew smaller, their protoplasm disappeared, and
they at last disintegrated. Pieces of the gall duct behaved differently.
They sometimes showed active growth, leading to the development of
numerous branched canals.[86]
Pieces of the kidney, when transplanted, suffered a great change, and
were subsequently absorbed. Transplanted pieces of a testis also
changed. After six days, Sertoli’s cells and the spermatozoa
disappeared. A kind of indifferent cell remained, characterized by clear
protoplasm and by a large nucleus. After seventeen days further changes
were observed, and later the pieces were completely absorbed. Pieces of
the ovary rapidly disappeared, leaving only a mass of interstitial
connective tissue.
The connective tissue underwent, in all the transplanted pieces,
characteristic changes. The tissue became less dense, the protoplasm and
nucleus of each cell enlarged. The cells multiplied, but only very
slowly. These changes took place after one or two days. After a month or
two the cells became more compact, their processes more numerous, and
the nucleus small and long. Later degeneration set in.
Small pieces of bone from the caudal vertebræ were also transplanted,
care being taken that each piece should contain some of the periosteum
and marrow. The bone tissue goes to pieces, but the periosteum and
marrow develop further. New bone is formed from the cells of the marrow
as well as from those of the periosteum. Finally the entire piece, both
its old and its new parts, is absorbed. Pieces of muscles were also
absorbed.
These experiments of Ribbert show that transplanted pieces of tissue do
not increase in size by growth, but undergo changes which he describes
as a return to an earlier condition of development. The abnormal
condition of their existence seems to be the cause of this change. The
transformation may be due to a change of nourishment, or to a loss of
nerve influence, or to lessened functional activity.
These results have a direct bearing on the problem of regeneration. They
show that all kinds of tissue may continue to live, and the cells
multiply in different parts of the body, but there seems to be nothing
in these cases comparable to a regeneration of the entire organ. In the
new situation the cells often assume an entirely new arrangement. After
a period of activity, a process of degeneration commences, and the piece
atrophies. Ribbert thinks that the atrophy is due to lack of
nourishment, yet it is not clear how this could be the case, since for
the first few weeks after transplantation there is an active growth, and
in some cases, as in that of the bone, there is a formation of new,
characteristic tissue. It may be that the transplanted tissues can no
longer manufacture the substances necessary for their specific growth,
and after the materials that have been brought along with them have been
used up, the growth of the piece is stopped and its subsequent
degeneration begins. It would be interesting to see if pieces
transplanted to the same kind of organ as that to which they belong will
become permanently incorporated in their new position.
The grafting-experiments that have been described in the preceding pages
were carried out with pieces of adult organisms. Somewhat different
conditions are present when parts of the developing egg or embryo are
united, inasmuch as a process has been started in them that may go on
independently, to a certain extent, of the union of the pieces. Born has
carried out a large number of experiments in grafting parts of tadpoles
of the same species, and also of different species. The union is brought
about at the time when the tadpoles are about to leave the jelly
membranes. The cut-surfaces are brought in contact and the pieces pushed
together and held in place for an hour or two by means of small silver
blocks or pieces of wire. The pieces readily stick together, and the
union is a permanent one. Before describing Born’s results, it may be
well to consider the power of regeneration of young tadpoles. If the
tail is cut off a new one is regenerated by the tadpole, but all parts
of the body do not have this same power. Schaper found that if a part of
the brain, or even the entire brain, is removed, no regeneration takes
place. I have found that if the region where the heart is about to
develop is cut out from a young embryo, a new heart is not formed.[87]
If a tadpole is cut in two across the middle of the body, neither piece
regenerates the missing half. Byrnes has found, however, that if the
region from which the posterior limb develops is cut out a new limb
regenerates. In older tadpoles, Spallanzani found that if the hind limb
is cut off it will regenerate, and Barfurth has more recently confirmed
this result. The end of the tail that has been cut off from a young
tadpole, before the tail has begun to differentiate, may continue alive
for several days. It grows larger and flatter, and the V-shaped
mesoblastic somites are formed. A slight regeneration even starts at its
anterior end, as first observed by Vulpian and later by Born. The
notochord and nerve-cord may send new tissue into the new part, and even
some of the muscle cells may extend into this part, but the piece dies
before regeneration goes any further. If, however, the tail is grafted
in a reverse direction on the body of another tadpole, the regeneration
may go further and produce a _tail-like_ structure, as Harrison
discovered and as I have also seen.
Born found that if the anterior half of one tadpole was united to the
posterior half of the same or of another tadpole a single individual was
formed which he kept alive in several cases until the time of
metamorphosis. If the head of a tadpole is cut off and grafted upon the
side of the body of another tadpole, the head will remain alive and
continue to develop in its new position, and, if well nourished by means
of the connecting blood vessels that develop, it may grow to be as large
as the head of the tadpole to which it is attached. Similarly, if the
tail of one tadpole is grafted upon the side of the body of another
tadpole, it also continues to develop, and at the time of metamorphosis,
when the normal tail is absorbed, the additional or misplaced tail also
shows signs of breaking down. Even the posterior half of one tadpole, if
grafted to the ventral side of another, may continue to develop,
producing legs, etc.
Born succeeded in uniting tadpoles of different species in several
different ways. They were united by their heads or by their ventral
surfaces, or longer and shorter tadpoles made by using pieces longer or
shorter than a half. In all of these cases there is no regeneration at
the place of union, and the internal organ, the digestive tract, nervous
system, and blood vessels unite when brought into contact. When pieces
are united end to end, like organs unite to like, the nerve-cord with
the nerve-cord, digestive tract with digestive tract, segmental duct
with segmental duct, cœlom with cœlom, and although less often, the
notochords sometimes join together. The lack of union of the ends of the
notochord is explained by its frequent partial displacement at the
cut-end, for when the cut is made the notochord, being tougher than the
other structures, is often dragged out of place in one or in both
pieces, so that the ends do not meet when the pieces are put together.
When like organs are brought together the substance of one unites
directly with the substance of the other, and if the organ is a hollow
one, as is the digestive tract or the nerve-cord, their cavities also
become continuous. There is also, Born states, some evidence to show
that if similar organs are not brought exactly in contact their ends
find each other and unite, and if they do not at first meet squarely
they may do so later. When the ends of unlike organs are brought in
contact, as, for instance, the nerve-cord and notochord, they do not
unite, but connective tissue develops between them. The union of like
parts, Born suggests, may be due to some sort of cytotropism, the
outcome of a mutual attraction between similar cells like that which
Roux has observed between the isolated cells of the segmented egg of the
frog. Born thinks that the first rapid union of the pieces is due to the
attraction of the ectoderm of one component for that of the other.
Born succeeded also in uniting pieces of the tadpoles of different
[Illustration: FIG. 54.--_A._ After Harrison. Union of two tadpoles by
posterior ends. Two days after operation. The line to the left of plane
of union indicates where the two were cut apart. _B._ Tail of right-hand
tadpole in _A_. Five days after cutting apart. _C._ Same. Nine days
after cutting apart. _D._ Same. Ninety-five days after cutting apart.
_E._ After Born. Combination of _Rana esculenta_ (anterior) and _Rana
arvalis_ (posterior). Thirteen days after the operation.]
species, even when they belonged to different genera. It is found,
however, that some of these combinations can be more easily made than
others, but it is not clear whether the difference depends upon
differences in the sizes of the pieces, or the rate of growth of the
ectoderm over the cut-surfaces, or to a deeper-lying lack of affinity
between the tissues. A combination of _Rana esculenta_ (anterior) with
_Bombinator igneus_ (posterior) was made. The combination lived for ten
days, and then showing pathological changes, it was killed. Another
combination is shown in Fig. 54, _E_, in which the anterior part of
_Rana esculenta_ was united to the posterior part of _Rana
arvalis_.[88] The blood of the posterior component was driven through
the vessels by the action of the heart of the anterior component. The
animal lived for seventeen days.
In all these combinations between different species, each developing
part retains its specific characters, and, although in several cases one
part received its nourishment from the other through the common
circulation, yet no influence of one component on the other could be
observed.
Harrison has succeeded in keeping an individual made up of two species,
_Rana virescens_ and _Rana palustris_, for a much longer time,--until,
in fact, the transformation of a tadpole into a frog had taken place.
Each half retained the characteristic features of the species to which
it belongs.
The absence of regeneration after the union of the pieces may be
attributed, in several cases, to the absence of this power in the region
through which the cut has been made; but in other experiments this
cannot be the explanation, since the power to regenerate can be shown to
exist in the part. This is the case in an experiment carried out by
Harrison and repeated later by myself. If the tips of the tail of two
tadpoles are cut off and interchanged (Fig. 55, _A_, _B_), a perfect
union takes place between the two parts, and a single tail develops.
Each of the cut-surfaces has the power to regenerate, but the union of
the parts has suppressed the regeneration. If, however, like parts are
not brought in contact, regeneration may take place in the region of
union (Fig. 55, _D_).
Both Harrison and I have made a number of experiments, in which the end
of the tail of a tadpole of one species was interchanged with a similar
part of another species. It is found that as the new tail grows larger
the ectoderm of the grafted piece is carried out to the tip of the new
tail, as shown in Fig. 55, _C_, and does not cover all the inner tissues
that belong to the same piece, the rest of the tail being covered by the
ectoderm of the major component. If the tip of the tail is now cut off,
as indicated by the line _b-b_ in Fig. 55, _C_, there are left at the
exposed edge two kinds of ectoderm, and from the cut-edge a new tail
regenerates, covered in part by each of the two kinds of ectoderm. I
made this experiment in order to see if the new ectoderm would show any
influence of its dual origin, especially along the line where the two
kinds are in contact, but no influence could be detected. In another
series of experiments the grafted tail was cut off, as shown in Fig. 55,
_A_, or in Fig. 55, _B_, or in Fig. 55, _C, a-a_. In these cases there
is left exposed, at the cut-edge, the internal tissues of the two
species. The new tail that regenerates is composed in part of material
derived from one species and in part from that of the other, but each
tissue remains true to its kind, and there is found no evidence of an
influence of one on the other (Fig. 55, _E_). These experiments show
that even when the two kinds of tissue regenerate side by side, and
unite to form a single morphological organ, there is no influence of a
specific kind of one tissue on the other.
[Illustration: FIG. 55.--_A._ _Rana sylvatica_ with grafted tail of
_Rana palustris_. Line _a-a_ indicates where tail was cut off. _B._
_Rana palustris_ with grafted tail of _Rana sylvatica_. Line _a-a_
indicates where tail was cut off. _C._ Older stage of a graft like _B_.
Lines indicating two possible operations. _D._ Another individual with
two tails, one composed of both components. _E._ Later stage of last,
when tail was cut off at level _a-a_.]
Another series of experiments in grafting, similar to one of those made
by Joest and myself on the earthworm, has been made by Harrison on the
tadpole. I have also later made similar experiments. Two tadpoles are
united by their posterior ends, as shown in Fig. 54, _A_, and a day or
two after union one of the tails is cut off near the line of union.
There is thus left attached to the end of the tail of one tadpole a part
of the tail of the other united in a reverse direction, so that the
exposed cut-end is the anterior end of the small piece. There grows out
from this cut-end a structure that resembles a tail (Fig. 54, _B_, _C_,
_D_). It contains a continuation of the notochord and nerve-cord, that
taper in a characteristic way to the end of the new structure. The tail
is flat and has a central band of muscle tissue, and a dorsal and
ventral fin. The muscles of the normal tail have a characteristic
V-shaped arrangement with the apex of the V’s turned forward, but
unfortunately in the new tail the muscles are so irregular that it is
impossible to make out their arrangement (Fig. 54, _D_). If the new part
is in reality a tail, the V’s ought to stand in the same way as do those
in the major component, and opposed to the V’s on the part from which
the new material arises. If the new structure is not a tail at all, but
a new growth, or even a suppressed trunk, then the V’s should stand as
in the small part itself. It has not been possible as yet to obtain a
decisive case. Harrison obtained one case in which the arrangement of
the muscles in the new part seemed to be more as it should appear if the
new part is a heteromorphic tail (Fig. 54, _D_). Even if this could be
shown to be the case, it may be that under the conditions of the
experiment the arrangement of the muscles is determined by the use of
the tail, although this does not seem very probable. Harrison, after a
careful analysis of the question, left it undecided, but seemed more
inclined to the view that the result is due to the development of
something new rather than a heteromorphic growth. On the contrary I am
strongly inclined to believe that the latter is the true explanation. In
another way I have been able to bring about the development of the same
structure. A small triangular piece is cut from the upper part of the
tail, as indicated in Fig. 56, _A_, one point of the triangle passing
through the notochord, or even through the aorta. If the cut-surfaces
are kept apart for a few hours, until the exposed end has been covered
over by ectoderm, they may not unite afterward, and two exposed surfaces
are left,--one at the distal end of the base of the tail, and the other
at the proximal end of the outer part of the tail. The latter surface
corresponds to that in the grafting-experiment. Regeneration may take
place from the two surfaces; both new parts seem to be exactly alike,
and both resemble a regenerated tail. The one from the proximal surface
of the outer part of the tail contains a notochord, nerve-cord,
connective tissue, pigment cells, and muscle tissue (Fig. 56, _B_). The
arrangement of the muscle fibres is generally very irregular, and the
characteristic V-shaped arrangement cannot be detected.
In only a few cases have attempts been made to unite two eggs or two
very early embryos, although there are a few casual observations[89] in
which such a fusion has been observed. The problems that arise in
connection with the union of two eggs are full of interest. Each egg has
the power of producing an embryo of normal size. If two eggs are united
into one, will a single giant organism result, or two organisms? If the
former, we must suppose that a new organization is formed of double
size. Whether an upper limit of organization exists can only be
determined by such an experiment. If two fused organisms result from the
fusion of two eggs, it would show the structure of the egg is of such a
kind that two organizations cannot readjust themselves into a single one
of double size. Moreover, it is important to discover whether any
difference exists as to the stage of development at which the union is
brought about, for it is conceivable that while a rearrangement is
possible at one stage, it might not be at another.
[Illustration: FIG. 56.--_A._ Tadpole to show where the V-shaped piece
is cut from the tail. _B._ Later stage of same with a new tail-like
outgrowth from the anterior end of tail.]
It has been shown that two blastulæ of the sea-urchin can be united to
form a single embryo. I found (’95) that occasionally two blastulæ stick
together and fuse, so that a single sphere of double size is formed. As
a rule two gastrulæ and two more or less complete embryos develop from
each double blastula, but in a few cases I found that a single embryo
may be formed, that shows, however, traces of its double origin. Driesch
has more recently (1900) succeeded[90] in bringing about more readily a
union of two segmenting eggs or blastulæ, and obtained perfect single
individuals from two fused blastulæ. He finds that if the fusion takes
place at an early stage the resulting embryo is less likely to show its
double origin than when older blastula stages are united. Zur Strassen
has also observed giant embryos of ascaris that arise by a fusion of two
eggs. Loeb has found that the eggs of chætopterus, which can be made to
develop parthenogenetically in certain salt solutions, often stick
together and produce giant embryos.