AMER. ZOOL., 14:673-698 (1974).
The Cellular Basis for Colony Form in Podocoryne carnea
MAX BRAVERMAN
SYNOPSIS. During the past 10 years considerable data have appeared indicating that the
growth of hydroids is not, as was earlier believed, similar to the meristematic growth
of plants, but rather that the sites of cell proliferation are removed in space from
the sites of utilization, that cells migrate individually, actively as amoebocytes through
the epidermis or pawively as epitheliocytes carried along in the hydroplasm, to the
sites of utilization, and that considerable migration across the mesoglca occurs.
A model of hydroid morphogenesis and morphostasis based on this new information
provides a comprehensive picture, albeit one filled in with numerous assumptions, of
the manner in which colony form is produced as a consequence of cell proclivities.
To contend that colony form or pattern in hydroids has a cellular basis appears, at first, to be a considerably belated
affirmation of the cell theory. But in speaking of a cellular basis for colony form, I
mean more than that there is an organization of cellular constituents underlying
gross colony form. I mean, rather, that a
number of aspects of the shape of a colony
are a consequence of specific cellular characteristics such as adhesivity or migratory
tendencies.
In the same way that a complex algebra
can be constructed from about twelve
properties of numbers, so it appears that
Financial support for the studies which form
the basis of this model came from a NATO
Postdoctoral Fellowship and an NIH Postdoctoral
Fellowship, Grants GB 5021 and GM 13708 from
the National Science Foundation and the U. S.
Public Health Service, respectively, and Institutional Grant CA 5184 to Allegheny General
Hospital.
Valuable technical assistance was contributed to
these investigations by, among others, Patricia
Becker and Frank Greenlee. Their patience and
care in culturing. photographing, and sectioning
material were indispensable.
The initial computer studies which provided a
point of departure for the investigations reported
here were carried out in collaboration with S. Ulam
and Robert Schrandt at the United States Atomic
Energy Laboratories, Los Alamos, New Mexico.
The theoretical orientation of these studies derives
strongly from S. Ulam's published papers and
from discussions with David Hawkins and Tryggve
Gustafson.
The address of the author is: Ranchos de Taos
Pottery, Los Cordovas Route, Box 131, Taos, New
Mexico 87571.
the complex colony structure is a consequence of a small number of cellular properties and their interactions. Gustafson
and Wolpert (1963) made a similar contention regarding morphogenesis in sea
urchins: Variation in a small number of
cellular proclivities accounted for the sequence of changing forms characterizing
the early development of the pluteus. But
the sea urchin is quite complex compared
to the hydroid colony and is following an
ever-changing developmental program. The
hydroid, growing limitlessly, and with one
exception1 growing repetitively, appears to
have but a single program, integrating
morphogenesis and morphostasis, which it
follows continually. The program is, of
course, written in the genes and mediated
biochemically, but it appears to be effected
by a small number of alternative cell characteristics which themselves are instructed
by a very large repertoire of genie permutations.
It seems that there are but few gates between the biochemical level of cell organization and the level of form-influencing
cell characteristics. The broad enzymatic
spectrum of differentiating cells has but few
morphogenetic expressions. A pin-ball
game with but four exit gates is a good
analogy. The number of paths the ball
can take is very high, corresponding to the
broad spectrum of biochemical alternatives
673
1
A discontinuity appears in the repetitiveness of
colony growth with the appearance of the sexual
person in the colony.
674
MAX BRAVERMAN
of cells, but, in the end, the ball falls
through one of four gates. The complex
path travelled influences which gate the
ball falls through. But conversely, which
gate it passes through does not reflect the
full complexity of the path by which it
arrived there. So, too, a cell's biochemical
constituency may influence these cell characteristics as a consequence of which animal form results: a very broad spectrum
of biochemical alternatives finding expression in only a small number of cell
characteristics.
Because only a single developmental program recursively instructs the continuing
growth and continuing maintenance of a
hydroid colony, there is a good chance that
the process can be known exhaustively.
That is, the factors influencing each cellular decision can be known, and the gross
influence of all decisions and their interactions assessed. To that end I have been
investigating the cell behavior underlying colony growth in Podocoryne carnea.
My first investigations (Braverman and
Schrandt, 1966) dealt mainly with the
biometrics of the colony and resulted in a
good knowledge of the statistics of colony
growth: total growth rate, branching and
new hydranth frequency and distribution,
individual stolon growth, and the relative
rates of these growth phenomena to each
other. My investigations at the cell level
have been aimed at determining the cellular basis for these phenomena. Simultaneously information has come from a number
of other laboratories bearing on the cellular basis of colony growth in a number of
hydroids. Although much detailed information is yet lacking, the time seems right
to try to fit this growing body of information together into a comprehensive theory.
Brien's hypothesis of the cellular basis for
the immortal hydra's morphostasis (1951)
has served us well. It has vividly captured
many aspects of the dynamic cellular behavior of hydra in a useful and picturesque
paradigm. Further, it has well served as a
measure by which to assess new data. But
the information accumulating from studies
of other hydroids begins to show a broader,
richer, and more complex picture of cell
movements than that originally observed
in hydra, and some studies of hydra's cell
movements also suggest a more complex
series of migrations than was originally
envisioned.
The new view emerging from my studies,
those of Hale (1964), of Sears Crowell and
his students (1965) and of other workers
(vide sotlo) emphasizes that colony form
is not a direct result of the sites of cell
proliferation. That it is was first suggested
by Kiihn (1910) who anticipated by 50
years current interest in recursion functions. Cell proliferation occurs for the
larger part in older, non-growing parts of
the colony. Cells then migrate from the proliferation sites to replace other cell types:
expendables such as nematocysts and digestive glands, and cells which themselves
become migratory, thus contributing to
colony morphostasis. Alternatively cells
move to newly forming colony parts, providing for colony morphogenesis. These
migrations occur in two ways: actively, as
amoebocytes migrate through the epidermal tissues, or passively as epitheliocytes
move into the gastrovascular cavity, where
they are carried about in the hydroplasm
to arrive by chance at their final destination. There, increased cell stickiness (an
important moulder of form in echinoderms, too) captures the peripatetic epitheliocytes and they become part of the
growing region. Thus, three principles
emerge as important to form generation in
hydroids: • (i) proliferation at sites distant
from the site of utilization; (ii) migration
of cells as amoebocytes and epitheliocytes;
(iii) random migration and cell stickiness
combining to accumulate cells at growth
regions. These are not, of course, the only
principles or rules of cell associations important in hydroid morphogenesis, but, at
the present time, they are the clearest and
seem the most important. How these rules
and others like them can interact to produce form is not immediately apparent
without evaluating the effect of their interactions. This can best be done by writing
a computer program, such as Schrandt and
I did for the rules of colony growth at the
"organ" level, and running the program
075
T H E CELLULAR BASIS OF HYDROID FORM
to determine what the interaction of rules
produces. In our case the computer output
was a two-dimensional array representing
the colony network. But the exhaustive
and quantitative knowledge of cell behavior on which a cellular model must be
predicated is still to be acquired. As a first
approximation to evaluating the effects of
interaction I shall consider, here, cell migration pathways, the patterns and priorities of colony growth, cell proliferation and
the nature of growth regions, and finally
attempt to synthesize, from these data, a
composite flow chart following cells from
their sites of origin to their sites of utilization and discard, and indicating cell decision points.
Podocoryne carnea is found in the eastern and western Atlantic and the Mediterranean on the shells of snails inhabited
by hermit crabs, where it grows, as do the
other Hydractinidae, completely adherent
to the substrate. Single polyps, removed
from the crab-borne colonies, attach to
microscope slides in standing sea water in
about 2 days. Stolons grow out from the
initiating polyps. They branch and anastomose with each other, and eventually new
hydrartths form on the stolons (Fig. 1).
Thus, the "pattern" of the colony is determined by (i) the rate of extension of established stolons, (ii) the sites of branching
(that is, of new stolon formation), and
(ill) the location of new hydranths in the
colony (Fig. 2). After a time, which varies
FIG. 1. A young Podocoryne carnea colony growing on a microscope slide.
STOLON
FIG. 2. P. carnea grows by increasing stolon length
and adding new hydranths. These two parameters
are mediated by three variables: (i) stolon extension, (ii) stolon branching, (iii) new polyp formation. (From Braverman, 19716.)
according to water temperature and other
conditions, medusae bearing sexual zooids
appear at the center of the colony and
spread circumferentially; meanwhile the
colony continues to grow to the limits of
the substrate.
In this study I shall attempt to explicate
the cell behavior which results in the three
aspects of colony form elaborated above,
in slide born colonies. I have no doubts
that the same principles apply to colonies
on other substrates. The model, for simplicity, will be limited to the presexual
colony.
CELL MOVEMENTS
From polyp epidermis to polyp
gastrodermis
In 1930 Kanajew reported that when
hydra is lightly stained in any of a number
of vital dyes, then removed, the dye is
localized primarily in the epidermis. Subsequently the dye disappears from the epidermis and is seen in the gastrodermis of
the animal. In addition to the Pelmatohydra oligactis with which Kanajew
worked, this movement of dye was observed
in Clytia johnstoni by Hale (1964), in
Podocoryne carnea (Braverman, 1969),
and in Hydra viridis by Shostak (personal
communication) .
This movement of dye might indicate the
movement of cells from epidermis to gastrodermis, or it might indicate the death of
676
MAX BRAVERMAN
the cells taking up the dye and their subsequent ingestion by adjacent gastroderm, or
it might indicate that the dye, but not the
original cells into which it was taken up,
moves into the gastroderm. Presumably,
one of the latter two interpretations has
been implicitly accepted, since Kanajew's
observation, which otherwise requires consideration, has, to my knowledge, been
ignored. The recent corroboration by other
investigators, however, and Rose and Burnett's extended investigations (I968a,fo,
1970a, b) on the epidermal source of secretory cells of the gastroderm of hydra and
of Cordylophora, recommend the first, and
obvious, interpretation: The dye, proved
stably resident in gastroderm cells (Burnett
and Garofalo, 1960), is equally so in the
cells of the epidermis, and therefore, dye
movement reflects regular cell movement
from epidermis to gastrodermis.
Rose and Burnett's strongest evidence
for the epidermal origin of gastrodermal
glands comes from experiments with Cordylophora (19706). Hydranths were given
an extended incubation in radioactive
thymidine, then the epidermis and gastrodermis were separated and the radioactive
epidermis combined with unlabeled gastroderm. The recombinant polyps were fixed
at regular intervals, sectioned, and the
tissue sections coated with photographic
emulsion. The radioautographs, bearing
exposed grains over nuclei containing
radioactive polynucleotide, indicate the
movement and differentiation of cells
initially taking up the radioactivity, that
is, cells preparing to divide.
At 10 hr after recombination small basophilic cells of the gastroderm are labeled.
In the succeeding sampling periods (15,
24, and 48 hr) immature zymogen and
mucous cells are labeled, and finally both
mature mucous cells and zymogen cells are
labeled. Rose and Burnett (19706) conclude that the epidermal interstitial cell
moves across the mesoglea to become the
gastrodermal basal cell, which differentiates
into the secretory cells of the gastroderm.
That some thymidine incorporating cell
moves from epidermis to gastrodermis and
then forms the secretory cells is clear. That
that cell is an interstitial cell could have
been rigorously established by showing that
the only radioactive cells in the epidermis
immediately after the labeling pulse are
those recognized as interstitial cells. Unfortunately, no zero hour sample was reported in this paper.
From polyp gastrodermis to hydroplasm
and thence to other parts of the colony
Rose and Burnett (1970b) show that
some cells moving from the epidermis to
the gastrodermis differentiate there as secretory cells. Experiments by Hale (1964)
using Clytia johnsloni and by Braverman
(1969) with Podocoryne carnea indicate
that in these colonial hydroids epidermal
cells marked with vital dye move into the
gastrodermis, and from there, through the
hydroplasm to other parts of the colony.
Clytia is a calyptoblast in which polyps
regularly regress and are replaced. Podocoryne is a gymnoblast in which, once
formed, polyps endure for the life of the
colony. Movement of cells out of Clytia
polyps occurs only when those polyps regress. Movement from Podocoryne polyps
occurs continually. With this exception,
passive migration of cells in the hydroplasm is identical in the two species.
Vitally stained polyp gastroderm cells
move to four locations in growing colonies.
1) From the proximal gastrodermis to
the distal gastrodermis. In grafts of Podocoryne polyps consisting of dyed bottoms
and undyed tops, the dye, initially in the
proximal epidermis, finally locates in the
distal gastrodermis. In the reverse graft,
dye slowly disappears from the distal epidermis. In neither case is there any indication of distal or proximal movement of
dyed cells in the epidermis (Fig. 3)
(Braverman, 1969).
This distad movement of gastrodermal
cells in Tubularia stems has been described
by Steinberg (1954, 1955) to be one of the
first steps to occur during hydranth regeneration. Four to 8 hr after decapitation of
a hydranth there is a distinct shifting of
T H E CELLULAR BASIS OF HYDROID FORM
'•'• STAINED EPIDCmiS
'//
STAMED WSTMOERMIS
FIG. 3. Vital dye grafts of Podocoryne carnea indicate no proximal movement of the dye. Dye
moves from the epidermis to the gastrodermis, and
distally in the gastrodermis. (After Braverman,
1969.)
endodermal cells in a distal direction
within the hydrocaulus. It is this distad
FIG. 4. New hydranths, forming on colonies initiated with dyed polyps, contain dye in their gastrodermal cells. This is more noticeable in starved
colonies, but occurs in fed ones as well. Stipple
677
movement, no doubt, that is responsible
for the gastroderm denudation that occurs
in repeatedly regenerating Tubularia stems
(Tardent, 1963).
2) To the gastroderm of neiuly forming
polyps and to the distal gastroderm of
sexual zooids. If Podocoryne hydranths are
removed from a colony, lightly stained in
Nile blue sulfate for 24 hr, then placed on
slides, they will attach there and initiate
new stolon and hydranth growth. As the
dye disappears from the epidermis, it can
be seen simultaneously to appear in the
gastrodermis of newly forming polyps
(Fig. 4) (Braverman, 1969).
When entire 3-month-old colonies of
Podocoryne carnea are lightly stained in
Nile blue sulfate, the dye initially resides,
as it does in single polyps, in the epidermis.
Young polyps are the most heavily dyed.
indicates dyed epidermis; solid color shows dyed
gastrodermis; numbers are month, day. (From
Braverman, 1969.)
G78
MAX BRAVEKMAN
The largest polyps in the colony take up
but little dye; sexual zooids take up none
at all. After 3 weeks of culture in undyed
sea water, very little dye remains in the epidermis. Considerable dye is resident in the
distal gastrodermis of young polyps, old
polyps and sexual zooids (Fig. 5).
3) To the stolon. Goldizen (1966) describes that when vitally stained hydranths
of P. carnea are placed on a microscope
slide bearing a young colony of the same
clone, they attach to the slide, and stolons
which form anastomose with those of the
colony already established. Two days later
ONE MONTH OLD COLONY
\
•
THREE MONTH OLD COLONY
FIG. 5. The distribution of vital dye in 1- and 3month-old colonies immediately and 3 weeks after
a 24-hr immersion in dilute dye. Stipple indicates
dyed epidermis; solid color shows dyed gastrodermis. (From Braverraan, 1969.)
T H E CELLULAR BASIS OF HYDROID FORM
679
palely stained cells can be observed scattered throughout the stolon gastrodermis
of the host.
4) To the stolon tips. Hale (1964) carried out extensive staining experiments in
which a hydranth (or a short length of
stolon) on a small piece of colony was
vitally stained. He describes that dyed
gastrodermal cells were transported to all
regions of the stolon (as well as to the
other colony regions described above) but
especially to the region 1 to 5 mm behind
the growing tip.
5) Special considerations. The passive
movement of the gastroderm cells through
the hydroplasm was observed by Goldizen
(1966) in time lapse films of growing
stolons. He describes intact cells floating
along in the hydroplasm. These cells adhered to the gastroderm wall of the stolon,
displaced adjacent cells, and adhered to
the mesoglea. Hale (1964) makes virtually
the same observation of cells floating in
Clytia hydroplasm.
I have seen, in histological preparations,
numerous examples of cells floating in the
hydroplasm of Podocoryne. In fact, in
many colonies the gastroderm of the stolon
appears to be a fluid tissue consisting of
rounded nucleated cells floating among the
contents of the hydroplasm (Fig. 6). In the
majority of the colonies, the gastroderm
was a discrete tissue of cuboidal cells,
clearly defined and firmly adherent to the
mesoglea. The liquid tissue rarely encompassed all the gastroderm of the colony.
Vitally dyed epidermal cells were never
observed to migrate directly to other regions of the colony without first passing
through the gastroderm. Apparently, the
cells incorporating dye move on a one-way
street from epidermis to gastrodermis and
thence to the hydroplasm for distribution
to other regions of the colony.
FIG. 6. The gastrodermis of P. carnea is a highly
labile tissue, a and b, Gastrodermal cells containing
nutriment float in the hydroplasm of the stolon.
(Arrows indicate the nuclei of these epitheliocytes.)
c, Zymogen-like cells (arrows) form in the stolon.
These are being carried in the hydroplasm. d, At
times the gastrodermis of the stolon appears
to be a fluid tissue. This is a newly formed
stolon. The bar, 50 p. in length, applies to all four
photomicrographs.
680
MAX
From stolon epidermis to hydranths
In older regions of the stolons of Podocoryne carnea, the epidermis is expanded
beyond the single layer found in the newly
forming stolon. These expanded regions
consist of cells with small nuclei, frequently bearing numerous division figures,
nematocytes in various stages of development, and other cells which are amoeboid
in shape (Fig. 7). The regions in which
nematocytes develop have been called cnidogenic regions (Bouillon, 1968) and have
long been known (Agassiz, 1862) . It is, of
course, not possible, looking at an amoeboid cell, to establish that it is a unique
cell type and not a cell on its way to becoming a cnidocyte. It is only the occurrence of these amoeboid cells in the upper
half of the polyp epidermis, and the rare
specimens caught in histological preparations half way between epidermis and gastrodermis, that indicate that under some
circumstances amoebocytes retain their
amoeboid shape until they reach their
FIG. 7. A, Amoebocytes, a, and B, cnidocytes, c, in
proliferation regions of the stolon of P. carnea. The
bar is 50 p. long.
ultimate tissue destination. The movement
of thymidine labeled cells, in colonies, further corroborates that many of the amoebocytes seen in amoebogenic regions of the
stolon (that is, those proliferative regions
giving rise to migratory amoebocytes) are
not in the process of differentiating as
nematocytes.
One-month-old colonies of Podocoryne
carnea are incubated for 1 hr in tritiated
thymidine. Radioautographs of tissue sections prepared from hydranths removed
from the colonies immediately after the
labeling pulse show which cells were in
the process of incorporating thymidine into
acid stable polynucleotide during the
pulse. The distribution of radioactivity in
hydranths removed from the same colony
24 hr later shows where the cells which
took up the label, originally, now reside.
Initially, virtually the only gastroderm
cells of the polyp to incorporate thymidine
are those of the hypostomal gland region.
(These are also the only gastroderm cells
to take up vital dyes.) Control experiments
investigating the incorporation of radioactive uridine and radioactive amino-acids
show that these substances are taken up
ubiquitously, not only by the cells of the
polyp, but by the cells of the stolon as well.
This guarantees that the distribution of
radioactive cells indicates the selective
incorporation, not exposure to, nor tissue
distribution of, radioactive thymidine.
In the epidermis radioactive cells are
found primarily in the mid-region of the
polyp, few above or below this. Considerable cell migration is indicated by the distribution of radioactive cells 24 hr after
the 1-hr labeling pulse. Now the entire
proximal polyp, both epidermis and gastrodermis, is heavily populated with radioactive cells (Fig. 8). Virtually no cells of
the proximal gastrodermis were initially
labeled; after 24 hr about 12% of the cells
were.
Conceivably, some of the increase in
labeled cells might be due to cell division.
Such increase, would then be proportional
to the original labeling. Surely this is the
case in the gastroderm of the upper third
of the polyp (Braverman, 1968, describes
681
T H E CELLULAR BASIS OF HYDROID FORM
THYMIDINE
Podocoryne
THYMIDINE
INCORPORATION
MITOSES
Tubuloria
Hydroctinto
24 HRS.
EPID.
GAST.
DISTANCE FROM
ORAL END
0
HRS.
DISTANCE FROM
ORAL END
24
HRS.
FIG. 8. Thymidine is incorporated only in the
mid-body epidermis and in the proliferating cells
of the hypostomal mucous gland region. Twenty-
four hours after a 1-hr pulse, both gastrodermal
cells and epidermal cells of the proximal polyp
are radioactive.
the cell movements in this apparently circumscribed polyp region). The increase in
proximal gastroderm and epiderm label is,
however, not proportional to the original
label and must therefore reflect the movement to this region of cells which were
resident elsewhere during the time of the
pulse. The only possible source for such
cells is the stolon.
Large numbers of labeled cells are found
in the cnidogenic and amoebogenic regions
of the stolon after a 1-hr tritiated thymidine pulse. For the most part, the labeled
cells are adjacent to concentrations of the
smaller cnidocytes and amoebocytes, and
resemble most characteristic cells of the
stolon. Thirty-five hours after such a pulse
considerably more cnidocytes and amoebocytes are labeled (Fig. 9). The large numbers of labeled cells in these epidermal
682
MAX BRAVERMAN
FIG. 9. Stolon proliferating regions (araoebogenic
or cnidogcnic) immediately after a 1-hr pulse
(a and a') and 60 hr after a 1-hr pulse (b and b')
with tritiated thymidine. In a and b the camera
was focused on the tissue section; in a' and b'
on the film emulsion. Only large cells are labeled
just after the pulse, but 60 hr later many small
nuclei are radioactive. This shows that the amoebocytes and cnidocytes are not a self-reproducing
population, but rather derive from the characteristic epidermal cells of the stolon. The bar is 50 /i
long.
proliferation centers indicates that these
could be the sources for the labeled cells
which migrate into the polyp. Unfortunately the polyp epidermis is so crowded
with small cells that a direct identification
of which cell type migrates into the epidermis is impossible. If the labeling pulse
is sufficiently concentrated and the exposure interval sufficiently long to provide
unequivocal labeling, then the grains mask
the morphology of the tiny cells over which
they lie.
The distribution of label in large nuclei
immediately after a labeling pulse, and in
small ones 35 hr later, indicates that
amoebocytes and cnidocytes are not a selfreproducing population, but rather arise
from characteristic cells of the stolon
epidermis.
Hale (1964) has described that no mitotic figures can be found in the gastroderm of either stolon or polyp of Clytia.
Nor have I been able to find more than a
few mitotic figures in the gastrodermis of
Podocoryne cornea, with the exception of
the circumscribed proliferation region of
the hypostome glands. Thymidine is, however, incorporated by the gastrodermal
cells of the stolon. This in itself is a puzzling situation. How can cells be continually incorporating thymidine into polynucleotide but never undergo division?
The entire question of cell division in
hydroids is puzzling. Podocoryne and
Clytia, at least, have very little Feulgendemonstrable DNA. Even in the epidermis
of Podocoryne very few division figures can
be recognized. Furthermore, multiple-
T H E CELLULAR BASIS OF HYDROID FORM
FIG. 10. Polynuclear cells (arrows) in the proximal
gastroderm of Podocoryne hydranths. These colonies were starved for 4 days, fed once, then starved
for 3 days more, m = mesoglea; g = gastroderm;
e = epiderm. The bar is 50 p long.
nucleated cells are frequently seen in the
gastroderm of Podocoryne (Fig. 10). Earlier investigators speculated that amitotic
division occurred in the hydrozoa (cf.
Braverman, 1971b). That is unlikely, but
the problem remains.
Since thymidine incorporation does occur in the stolon, stolon gastrodermal cells
could be responsible for the increase in
labeled cells of the polyp gastrodermis.
Gastrodermal cells, after synthesizing DNA,
might round up and lose their connection
to the stolon mesoglea to float in the
hydroplasm. It is possible that these cells
might then insinuate themselves among
the proximal polyp gastroderm but it is
unlikely, for cell movement in the polyp
gastroderm is from proximal to distal, suggesting that the distal region is stickier
than the proximal; furthermore, dyelabeled cells moving into polyps tend to
become established in the distal, not the
proximal region. It is more likely that the
source of labeled cells in the polyp epidermis and gastrodermis is the amoebocytic
cells which migrate up into the polyp
from the stolon, some of which move across
the mesoglea into the gastroderm.
That similar cells migrate in the epidermis of other species has been previously
reported by Tardent and Morgenthaler
(1966) and by Lenhoff (1959). The latter
investigator demonstrated the distal migration of 14C labeled cnidocytes. Burnett
683
et al. (1968) have described how in tissue
cultures of Tubularia cells, first endoderm
cells migrate out from the inoculum, then
cells they describe as interstitial cells follow. In Cordylophora, similar cells move
across the mesoglea and differentiate as
gland cells (Rose and Burnett, 197O£>).
Indeed these migrating amoebocytes appear to bear many of the properties previously claimed for the interstitial cells, with
the major exception that they are not a
self-sustaining population but arise from
epidermal cells of the stolon.
In summary, then, the change in distribution of labeled cells which occurs
between 1 and 24 hr after the labeling
pulse is due primarily to the migration of
amoebocytes from stolon proliferation
regions into the polyp epidermis and across
the mesoglea into the gastrodermis.
Out of the stolon gastrodermis
The gastrodermis of the stolon of Podocoryne carnea varies in its structure. At
times it can be seen to be a discrete, regular tissue bearing regularly shaped gastrodermal cells which entirely cover the mesogleal surface; at other times, the tissue
appears virtually fluid: cells are not adherent to the mesoglea but rounded up,
packed tightly in some areas of the stolon,
absent from others (Fig. 6). This apparent fluidity is consistent with the picture
of gastroderm fluidity described by Steinberg (1954, 1955) to occur during Tubularia regeneration and by Mookerjee and
Bhattacherjee (1966) to occur during
hydra regeneration. Burnett (personal
communication) has also described gastrodermal cells of the hydra polyp breaking
loose and floating about in the gastrovascular cavity.
It seems that under circumstances at
present not clearly understood, but certainly during starvation and during polyp
regression, but also under more normal
circumstances, cells of the gastroderm lose
their adhesiveness, floating free from the
mesoglea to move wherever the currents
and pulsations of the hydroplasm carry
684
MAX BRAVERMAN
them. In those hydroids undergoing regression and replacement of their polyps
(probably calyptoblasts in general)
(Crowell, 1953) gastroderm release occurs
solely during polyp resorption. In species
in which polyps are not regularly resorbed
(probably gymnoblasts in general) release
of gastroderm cells occurs during normal
colony growth. In both, gastroderm cells
seem to be continuously attaching and
releasing from the stolon.
Summary of cell movements
not to others, that nutrition is equitably
distributed among colony regions. Stolon
growth (the parameter most sensitive to
starvation in Podocoryne) proceeds in
starved halves of colonies at the same rate
as in the halves which receive food (Braverman, 1971a). Thus, the mixing of nutriment in the stolon network and/or the
indirect influence of nutriment on growth
is complete and equitable. The proximity
of stolon regions to nutriment sources, the
feeding polyps, plays no role in morphogenesis.
Colony starvation does, however, differentially affect the growth parameters. This
will be considered below under Priorities.
Two major types of cell movements are
found in the hydrozoa. Epidermal cells migrate as amoebocytes and cnidocytes from
their proliferation sources in the stolon Temporal constraints
and in the mid-regions of polyps into
Important temporal constraints limit the
polyps and thence across the mesoglea into
the gastroderm, or up into the tentacles. morphogenic capabilities of the stolons.
Gastrodermal cell movements are passive Hydranths form on stolons of 2, 3, and 4
and involve a loss of adhesivity of gastro- clays of age. Twice as many form on 3- and
dermal cells to the underlying mesoglea 4-day-old stolons, and about as many form
and release to the hydroplasm. There gas- on 2-day-old stolons as random distribution
trodermal cells are carried about, passively would predict (Braverman, ]971«).
propelled by currents created by hydroBranches tend to form on somewhat
static pressures of polyps filling and empty- younger stolons than do new hydranths.
ing, by peristaltic contractions of the sto- Two and a half times the randomly prelonic contractile zone, and by the flagella dicted number form on 2-day-old stolons;
of the cells of the gastroderm.
1.3 times the predicted number form on
At this time it is not at all clear whether 3-day-old stolons (unpublished). Cnidothe sole proliferating tissue in hydrozoan genic and amoebogenic regions tend, at
colonies is the epidermis (with the ex- least in younger parts of the colony, to be
ception of the circumscribed proliferating located at branch points. The physiologiregion of the hypostomal gastroderm), or cal basis for these temporal constraints is
whether the thymidine-incorporating cells presently unknown.
of the stolon gastroderm function as a cell
source despite the lack of noticeable mi- Feedback regulation
totic figures in this tissue. A clear picture
Hydranth formation is further conof cell movements and cell proliferation is strained as a feedback function apparently
dependent upon the resolution of this related to the number of hydranths present
paradoxical situation regarding gastroderm on a colony. In those colony halves from
cell proliferation.
which polyps were removed, about two and
a half times as many hydranths formed as
PATTERNS AND PRIORITIES OF
formed in the control halves during the
COLONY GROWTH
same time (Braverman, 1971a). Thus,
there is some kind of control which reguNutrition
lates the number and location of new
Both Crowell (1957) and Braverman hydranth formation. Since the control
(1971a) have shown, by selectively provid- applies to where hydranths form, it is uning food to some regions of a colony and likely that materials moving in the hydro-
685
T H E CELLULAR BASIS OF HYDROID FORM
ber of new branches relative to the amount
of new stolon growth (Table 1). Those
individual stolons already in existence continue to grow at the same rate as their
counterparts in fed controls. This is dramatically demonstrated by photographs of
colonies that were starved for 9 days, beginning on the 20th day of culture, compared to controls that were fed during the
entire time. The peripheral area subtended by the stolons of the starved colo-
O
DC
100
80
O 60
o
LJ 20
Q.
DAYS
FIG. II. The rate of stolon growth falls off linearly
and immediately in starved colonies of P. carnea.
Hydranth formation remains at the control level
for 2 clays. (From Braverman, 1971a.)
plasm exert this control, as they, if food
distribution be any criterion, are ubiquitously distributed.
Priorities
Although the rate of growth of hydroids
is known to vary with the nutrient input
(cf. Davis, 1971), starvation affects the
growth variables of the colony in different
ways. Thus, by starving mature colonies
the natural growth hierarchy can be demonstrated. When colonies of Podocoryne
are starved, stolon growth falls off linearly
and immediately; hydranth formation lags
2 days in demonstrating the effects of starvation (Fig. 11). The total length of stolon
growth in starving colonies is reduced, but
the growth of individual stolons is not.
Frequency distribution analysis (Fig. 12)
of the lengths individual stolons grow in
starved colonies and in fed controls shows
that those stolons which do grow in starved
colonies grow the same amount as those
growing in fed controls.
The difference in total stolon length
grown reflects the difference in branching.
In starved colonies, branching is severely
curtailed. The absolute number of new
branches is reduced, as well as is the num-
4
6
8 10 12
14 16 18 20
STOLON LENGTHS
Class mid marks
FIG. 12. Frequency distribution of the lengths individual stolons grow in colonies starved for 4 days
(A) a n c l ' n control colonies (%) fed daily. Measurements were taken on each of 4 days and
summed. The colonies were photographed daily.
Each day a colony was traced onto an overlay
from a 15x enlargement. Thereafter each day's
growth was recorded and added to the overlay.
n fed = 1148; n starved = 698. The rate of growth
of already existing stolons in starved colonies does
not differ from the growth of stolons in control
fed colonies.
686
MAX BRAVERMAN
TAI'.LE 1. Branching and .stolon growth.
New stolon
Fed
New hydranths
New branches
New branches/
New hydranths/
New stolon length New stolon length
(X 100)
(X 100)
Starved
Fed
Starved
Fed
Starved
Fed
Starved
Fed
Starved
Day 1
2
3
4
235.9
283.4
331.7
392.3
221.3
193.5
162.6
102.4
3.5
8.5
8.9
4.8
4.1
6.6
2.4
1.4
12.4
18.1
27.0
30.4
5.6
5.0
5.2
1.9
1.5
3.0
2.7
1.2
1.8
3.4
1.5
1.4
5.3
6.4
8.1
7.7
2.5
2.6
3.2
1.8
Total
1,243.3
679.8
25.7
14.5
87.9
17.7
8.4
8.1
27.5
10.1
nies is equal to that of the fed colonies, but
the stolon density is considerably less
(Fig. 13).
Although "new branches/new stolon
length" was reduced in the starved colonies to about y3 the number of those in
the fed, the number of "new hydranths/
new stolon length" was the same in starved
and fed colonies. From these data a morphogenetic priority can be constructed.
Stolon extension will continue despite
starvation; new hydranth formation will
occur on stolons of the appropriate age,
and according to some feedback regula-
FIG. 13. Colonies fed (right; and starved (lett)
for 1 week beginning on their 20th day. The
colonies are depicited on day 20 and day 27. The
stolons of the starved colony extend as far periph-
erally as do those of the fed colony, but branching
in the starved colony has been severely repressed.
The bar is 1 cm long.
THE CELLULAR BASIS OF HYDROID FORM
tion, not at the present time understood,
in proportion to the length of new stolon
formed. Branch formation, however, will
be curtailed in response to limited
nutrition.
Similar results regarding growth priorities were reported by Crowell (1957) on
the basis of experiments similar to these.
Crowell observed, ". . . there is good correspondence between the total quantity of
the stolonic system and the quantity of
food," but he showed that branching to
form uprights is severely restricted under
starvation conditions. This is true, he reports, in both the calyptoblast Campanularia flexuosa and the gymnoblast Cordylophora sp. Crowell's analysis of growth
priorities, taking into account an earlier
description by Berrill (1950), is lucid: "It
seems to be a clear rule that locations
where proliferation is occurring have
precedence over zones of prospective
growth." This was written before Crowell's
vital dye studies led him to question that
growth regions are proliferation regions
(Crowell et al., 1965).
Growing stolon tips are characterized by
a cap of epidermal tissue, elongated in the
direction of stolon growth. These cells
bear what appear to be vacuoles proximally. The vacuoles are adjacent to other
vacuole-like structures at the distal border
of the gastrodermal cells. Whether these
are intra-cellular vacuoles or simply empty
spaces is not clear to me. The nuclei of the
tin epidermis characteristically bear proximally directed adherent chromatic appendages. Perhaps these are associated with their
perisarc depositing function. The structure
of the tip gastroderm can be either well or
poorly denned.
By means of time lapse cinematography,
Goldizen (1966) demonstrated a pulsating
pattern of advancement of the tip of Podocoryne similar to that shown for Clytia
(Hale, 1964) and for Campanularia (Wyttenbach, 1968). There appears to be some
correlation between the extension and retraction of the tip, the degree of fluidity
of the gastroderm, and the distribution of
a bolus of cells and free nuclei which is
frequently seen during the contraction
phase, some distance behind the tip. Although it is difficult to be certain that the
effect is not an artifact of tangential section, it also appears that during retraction
the epidermis contains an increased number of the aforementioned vacuole-like
spaces (Fig. 14).
The pattern of movement of vital dyes
FIG. 14. The gastroderm is of the stolon tip varies
from a discrete, regular tissue (a) to a poorly
defined, virtually liquid tissue (c) . This change in
tissue fluidity may lie related to stolon extension
and the pulsating extension and retraction of the
tip. The bar is 50 n long.
THE NATURE OF GROWTH REGIONS
The stolon tip
688
MAX BRAVERMAN
•NEW STOLON-
-TIP*to
£0<
o
cc
15
10
LU
.0-
_L
.05
.10
.15
_L
.20
25
.30
DISTANCE FROM TIP
FIG. 15. T h e distribution of radioactive nuclei in
the stolon tip immediately after (a and c) and 35
h r after (b) a 1-hr incubation in tritiated thymidine. Although tip nuclei do not incorporate thyrnidine, labeled nuclei are present at the tip 35 h r
.35
.40
45
.50
(mm.)
later. A possible explanation is that epitheliocytes
float through the hydroplasm, adhere to the tip
a n d insinuate themselves among the cells of both
the epidermis and gastrodermis. T h e bars are 50 p
long. (In part, from Braverman, 19716.)
THE CELLULAR BASIS OF HYDROID FORM
689
in the stolon, relative to the movement of
the elongating tip (Overton, 1963; Hale,
1964; Crowell et al., 1965), indicated that,
contrary to earlier expectation, stolon extension was not effected by terminal proliferation in a meristem-like region. The
indirect evidence of dye movements has
subsequently been directly verified. Little
thymidine incorporation occurs at the
stolon tip. The per cent of labeled cells
increases more or less regularly for the
first 0.5 mm from the tip. Incorporation
is approximately equally divided between
epidermal and gastrodermal cells (Braverman, 1971&). Since autoradiographs show
that uridine and amino-acids were incorporated ubiquitously by cells of the
stolon into acid stable material, the pattern
of thymidine incorporation can be taken
to indicate which cells are incorporating
that precursor.
Thirty-five hours after a 1-hr labeling
pulse with tritiated thymidine, however,
many labeled cells can be seen in both
epidermis and gastrodermis of the tip (Fig.
15). The gastrodermal cells no doubt
arrive there carried by the stolon currents.
It is not equally clear how labeled epidermal cells can be present at the tip unless (i) tip cells are regularly sloughed, or
(ii) gastroderm cells move into the tip
epidermis.
FIG. 16. Branch point genesis. The initial raorphological event consists of elongation of epidermal
cells in the direction of new stolon. The bar is
50 p long. (From Braverman, 1971 b.)
New branches
The sequence of events characterizing
branch formation begins with reorientation of epidermal cells perpendicularly to
the mesoglea. The cells continue to elongate. Their nuclei develop characteristic
proximally directed tabs. The perisarc is
dissolved and, shortly thereafter, gastrodermal cells reorient with respect to the
new tip. By histo-morphological criteria,
690
MAX BRAVERMAN
FIG. 17. Stolon anastomosis. Perisarc dissolution in
the recipient stolon is effected by branch formation
induced in that stolon by the anastomosing one.
A, Phase contrast photomicrograph of living
stolons. li-D, Stages in anastomosis. The bar is
125 /i long for A; 50 fi long for B-D. (From Braverman, 1971b.)
branch formation appears to be an event
initiated by the epidermis. No increased
incidence of thymidine incorporation is
present in the cells which reorient to form
new branches, thus direct proliferation is
no more responsible for new branch formation than it is for stolon extension
(Fig. 16).
Stolon anastomosis is, interestingly, accomplished via the same mechanism as
branch formation. As a stolon tip approaches another stolon, the cell morphology characteristic of the epidermis of
new branching is induced in the recipient
stolon. This morphogenetic induction at a
distance, across intervening sea water, is to
my knowledge a unique phenomenon (Fig.
17). It has previously been described to
occur in anastomosing stolons of Hydractinia echinnta (Miiller, 1964). Thus, apparently, is the perisarc broached.
Goldizen (1966) has described that advancing tips which approximate empty
stolons of old portions of the colony do
not invade the empty stolons, but move
over them, corroborating the observation
that perisarc dissolution is accomplished
by the recipient stolon. Subsequently the
two epidermal layers fuse, and, by a
mechanism unknown to me, the fused
epidermal layers part and the gastroderms
become confluent.
New polyps
Polyp formation in Podocoryne, unlike
that in hydroids with terminal polyps, requires that a new "growth" region be
initiated along with polyp differentiation.
I previously (Braverman, 1971/;) described
this new region as being identical in origin
to that of stolon outgrowth (branching).
THE CELLULAR BASIS OF HYDROID FORM
691
Two important aspects of the cellular
mechanisms relevant to the growth and
maintenance of hydroid colonies remain
yet unknown, (i) Gastrodermal cells incorporate thymidine but do not demonstrate mitosis. Do these cells divide? If so,
how? Alternatively, is the epidermis the
only proliferation source in the colony?
(ii) How does extension at the stolon tips
occur? Where do the epidermal and gastrodermal cell components for each of these
come from?
In order to construct a model based on
the source, movement and distribution of
cells in growing colonies I shall make the
following assumptions with regard to the
two problematical aspects of cell proliferation and growth.
1) Gastrodermal cells of the stolon can
give rise to other gastrodermal cells (no
assumption of method is necessary). This
applies only to gastrodermal cells of the
stolon; little thymidine incorporation is
demonstrated by gastrodermal cells of the
polyp stalk, so it is reasonable to assume
that although they cycle throughout the
colony as epitheliocytes, no cell division
occurs in the polyps.
2) Stolon tip extension results from division of cells distributed among the adjacent stolon epidermis and from the adhe-
FIG. 18. Three stages of development of a new
polyp. All are characterized by a bolus (B) of
cellular and nutritive material. E = epidermis;
G = gastrodermis; F = free cell; N = nematocyst;
T = tentacle. The bars are 50 ^ long. (In part,
from Braverman, 1971b) .
In that description I was mistaken, and
probably misled by aberrant stolon tips
growing upward from the surface of the
substrate. Although it remains difficult to
identify unequivocally the earliest stages
of hydranth outgrowth, the first recognizable form appears to be sphere shaped
and without perisarc (Fig- 18). Berrill
(1961) described that not only are what
appear to be vacuolated cells characteristic
of the advancing terminal stolon, but also
these vacuolated cells are present in terminal hydranths developing, in Obelia and
Campanularia and many related genera, at
the ends of stolons. These vacuole-like
structures seem to be ever present in hydroid regions of tissue extension. They may
well play some critical role in accommodating the cellular accretion which must
occur there.
A MODEL FOR THE CELLULAR BASIS OF
MORPHOGENESIS AND MORPHOSTASIS
IN HYDROIDS
MAX BRAVERMAN
692
EPITHELIOCYTES
STOLON
EXTENSION
CELL DIVISION
ANTERIOR TO
FIRST BRANCH
IF
Ke )
IF
K e = Kg
Kg
STOLON
EXTENSION
and
BRANCHING
STOLON
EXTENSION
EpWttliocytos
FIG. 19. The cellular basis of stolon extension in P. carnea.
sion to the growing region of epitheliocytes
floating in the hydroplasm. I shall further
assume that under conditions of starvation,
and perhaps under other conditions presently unknown, the contribution from
adjacent epidermis is minimal and that
the majority of cellular material for growth
is obtained from epitheliocytes.
Stolon extension and branching (Fig. 19)
Stolon extension in the epidermis of
Podocoryne carnea proceeds partially or
wholly by virtue of cell division occurring
between the stolon tip and the closest
stolon branch. The extension of the tip
gastrodermis is accomplished by epitheliocytes which float in the hydroplasm, accumulate at the stolon tip, and interdigitate among those already there, during the
extension phase of tip pulsation. Presumably, advancement of the two layers is
balanced by the extent of epitheliocyte
contribution to the epidermis. This is
maximal during periods of starvation. At
these times no cell division occurs in the
stolon epidermis; the only source of cells
for stolon extension is epitheliocytes released from the gastroderm of older parts
of the colony.
In a normally fed colony a proliferative
balance exists between the rate at which
the tip moves forward and the rate of proliferation in adjacent stolon epidermis. If
the latter outruns the former, the density
of cells in the space denned by the mesoglea and perisarc laterally, and the tip and
a branch antero-posteriorally, increases.
This increase in cell volume within a constrained space stimulates epidermal cells to
reorient in the characteristic palisaded
form of new stolon branches, and stolon
outgrowth occurs at that point. In starved
colonies no cell division occurs in the
distal epidermis and thus no branching
occurs.
Hydranth formation
Although some of the constraints on
hydranth origin are known, viz., (i) that
693
THE CELLULAR BASIS OF HYDROID FORM
hydranths form preferentially on stolons
of 2 and 3 days of age, and (ii) that some
feedback control regulates the distribution
of new hydranths, there is no evidence relating to the cellular basis of either of
these constraints. Thus, it does not now
seem that hydranth formation is a consequence of the modes of proliferation and
cell movement which characterize stolon
extension and stolon branching.
A simple mechanism for hydranth genesis, and one that is consistent with the
constraints demonstrated in P. carnea and
with what is known of cell movements
during Tubulnria regeneration, is the following. Inhibition of gastrodermal stickiness is effected by some substance of finite
lifetime which has its origin in hydranths
and moves through the cells of the adjacent stolon. When new stolons form sufficiently far from the existing hydranths,
and when the stolon tissues reach an age
which renders them competent, gastrodermal cells or their underlying mesoglea
RANDOM
MIGRATION
IN STOLON
T
become sticky, thus attracting a concentration of gastrodermal cells. Attractive as this
theory is, I have found no histological evidence to corroborate it, not, perhaps, because it does not occur, but rather because
small accumulations of gastrodermal cells,
in stolon still bearing perisarc, would be
indistinguishable from the usually observed
distribution of gastrodcrni which appears
very irregular.
Epidermal regions of localized proliferation
(Fig. 20)
Autoradiographs show that between 15
and 20% of the epidermal cells of the
stolon are synthesizing DNA during a 1-hr
period. In newly formed stolon distal to
the first branch this proliferation no doubt
contributes to stolon extension. Proximal
to the first branch, cnidogenic and amoebogenic regions are formed. Presumably, the
constant level of epidermal proliferation is
channeled into these compartments in
DISTAD
MIGRATION
IN POLYP
NEMATOCYTES
STOLON
PROLIFERATIVE
REGION
AMOEBOCYTES
T
RANDOM
MIGRATION
IN STOLON
ZYMOGEN
CELL
MUCOUS GLAND CELL
GASTRODERMAL
QGESTTVE CELL
FIG. 20. Proliferation and migration of amoebocytes and cnidocytes in P. carnea.
694
MAX BRAVERMAN
older regions of the colony. The specific
stimulus to this developmental rechanneling is presently unknown, but the appearance of amoebogenic and cnidogenic
regions at branch points is suggestive of sequential and cumulative differentiation.
Epidermal cells divide, and dividing become smaller. The daughter cells probably divide again. These cells become either
amoebocytes or cnidocytes. In either case
at some stage in their differentiation they
migrate from the proliferation region.
Their movements in the stolon itself are
most likely at random, but a constraint is
probably imposed which favors distal migration into and through the polyp. In the
polyp, the cnidocytes migrate to the tentacles; the amoebocytes, at each level, make
a decision whether to cross the mesoglea
or move upward. This decision is apparently influenced by the numbers of gastrodermal digestive cells and gastrodermal
gland cells at each level (Burnett, personal communication with respect to zymogen glands). How this influence is mediated is presently unknown.
Adjacent epidermal cells are continuously recruited into the proliferation
regions of the stolons, and probably into
that of the polyp too.
Gastrodermal digestive cells (Fig. 21)
Of the polyp. The factors regulating the
duration of residence of gastrodermal digestive cells in a polyp are presently unknown. On the one hand, thymidine incorporation does not occur in the polyp
gastroderm but does occur in gastrodermal
cells of the stolon, so that it is likely that
m <3UF£RATK)M
\
STOLON
OASTK00CMIIS
ZYMOGEN
GLJUO y
i
\
J
OASTDHESTIVE
CELLS OF POLYP
/
STOLON T I P |
digestive cells ingesting nutriment, in the
polyp, lose their attachment and migrate
to stolon regions where they initiate DNA
synthesis. On the other hand, the polyps
of starved colonies are divested of gastrodermal digestive cells. The continued
growth of stolon in starved colonies suggests that even during starvation epitheliocytes are available for stolon extension.
Thus, it appears that gastrodermal cell
release is effected at a constant rate, insensitive to nutritive conditions.
As with amoebocytes and cnidocytes,
some force stimulates gastrodermal cells
to move distally in polyps. This could be
effected by reducing adhesive forces proximally relative to those in the distal polyp
gastroderm. Conceivably changes in adhesivity, and the distal attraction of
amoebocytes and cnidocytes, might reflect
the differences in electrical potential which
Rose (1963fl,£>) has suggested play an important role in defining the circumstances
under which regeneration can occur in
Tubularia.
Whatever the regulative mechanism, digestive cells lose their contact with the
mesoglea and move into the hydroplasm
as epitheliocytes. There they are carried
passively until they come into contact with
a region that is sufficiently sticky. Such
sticky regions exist, apparently, at the stolon
tip, in newly forming hydranths, in the
distal portion of all hydranths, and in
some regions of the stolon.
Of the stolon. Gastrodermal digestive
cells of the stolon carry out polynucleotide
synthesis, presumably synthesizing DNA.
Presumably some form of proliferation occurs, despite repeated failure by a number
of investigators to identify mitotic figures
in cells of the stolon gastrodermis (Hargitt, 1903; Billard, 1904; Hale, 1964). Cells
are released from the stolon gastrodermis,
under circumstances presently not understood, into the hydroplasm, where they
join the epitheliocyte population.
EPITHELIOCrTEl^
l f ^ l MEW POLYP
On interstitial cells
CELLS
|
FIG. 21. Epitheliocyte origin and migration in
P. carnea.
In his book, The Germplasm, August
Weissmann (1892) spelled out his inter-
THE CELLULAR BASIS OF HYDROID FORM
pretation of the Roux-Weissmann hypothesis that cell division could distribute the
hereditary material equally according either
to quantitative or qualitative standards. He
hypothesized, "Ontogeny depends on a
gradual process of disintegration in the
development of each individual. . . . Finally, if we neglect possible complications,
only one kind of determinant remains in
each cell, viz. that which has to control
that particular cell or group of cells"
(quoted in Wilson, 1911).
If cells of the adult phenotype contain
but a small fraction of the hereditary
determiners, then it is necessary that a
special group of cells be set apart as germ
cells. Weissmann's ideas about the location and behavior of germ cells derive primarily from his study of hydroids, indeed,
from his study of Podocoryne carnea. How
Weissmann's theorizing distorted his observations is reviewed by Berrill and Liu
(1948). Similarly, the high degree of regeneration of which hydroids are capable
was inconsistent with the severely restricted
genie potency of adult cells, unless one
hypothesized that along with the germ
line a second group of cells, embryonic in
that they retained qualitatively complete
nuclei, was set aside. These were the interstitial cells which ostensibly provided totipotent nuclei for regeneration. Even in
1911 Wilson identified the "quasi-metaphysical character which almost places it
outside the sphere of legitimate scientific
hypothesis" of Weissmann's theorizing
(Wilson, 1911, p. 407). He cites, in contradiction, the general observation that in
ordinary mitosis, "the division of the chromatin is carried out with the most exact
equality," and cites also numerous experiments in which complete embryos resulted
from isolated blastomeres.
Experiments with hydroids by Zwilling
(1963), Normandin (1960), Haynes and
Burnett (1963), and Diehl (1968) leave
no doubt that interstitial cells are not
requisite to regeneration of whole animals.
A remnant of the Weissmannian view that
only I-cells are toti potent resides, nevertheless, in the belief that interstitial cells
are a self-sustaining population, and that
if they are selectively destroyed, they cannot be replaced from other cell types
(Brien and Reniers DeCoen, 1949; Diehl
and Burnett, 1964). Both experiments
upon which this conclusion rests depend
on an agent presumed to be selectively
effective against interstitial cells; in one
•Msry; > -•
FIG. 22. "Interstitial cells" (i) , top, and cnidocytes
(en), middle, in hydra. Bottom, cnidocytes (en)
and amoebocytes (a) in P. carnea. I-cells are ovoid
with a single nuclcolus; amoehocytes are amoeboid
and contain much dispersed chromatic material.
Both preparations are stained with toluidine blue.
Hydra cells are much larger than those of P. carnea.
The bar is 50 p. long.
MAX BRAVERMAN
696
constructed rigorously would be preferable.
Nevertheless, the construction of an explicit model from inadequate data has the
virtue of identifying clearly what is known,
making assumptions explicit, and emphasizing what is unknown. Attention is
thus focused on areas requiring further
investigation.
The model is a complex one. For example, although starvation can be seen to
reduce branching, it is not apparent what
effect starvation would have on the other
compartments of the model. Even if the
assumptions used to construct this model
prove to be false, it seems to me that working out the dynamics of interaction among
the different compartments is a worthwhile
exercise, if only to acquire expertise with
the kind of interacting system which is
likely to prove central to developmental
phenomena.
A glance at the model representation
(Fig. 23) shows that the idea of simple
causality has little meaning in evaluating
complex interacting systems. Indeed, each
of the diamond shaped decision points inSummary and conclusions
dicates a cellular decision with which a
Many assumptions have been utilized to "cause" can be associated, but in a more
construct this model. Naturally a model important sense that "cause" of cell dis-
case radiation, in the other nitrogen mustard. But that assumption, in fact, assumes
the conclusions, inasmuch as both are effective against any dividing cells. Thus, not
only I-cells but also any other cell which
might divide to give rise to I-cells would
be affected.
It has been necessary to consider the
nature of the interstitial cell and the controversy surrounding it, because in many
respects the amoebocytic cells of Podocoryne resemble what has been described
as interstitial cells. They are migratory,
differentiate into a number of cell types,
and reside in the epidermis. Amoebocytic
cells of Podocoryne do not morphologically
resemble, however, the nests of I-cells which
are present in Hydra (Fig. 22). Nevertheless it seems likely that the role of I-cells
and the role of amoebocytes is similar.
Neither uniquely totipotent nor a selfreproducing population, they are the proliferative and migratory phase of epidermal
cells of the hydrozoan.
;»|EPITHELIOCVTE.[
>
---» T
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Y1G. 23. The cellular basis of morphogenesis and morphostasis in Podocoryne earned.
THE CELLULAR BASIS OF HYDROID FORM
tribution in any compartment can only be
the nature of the entire system. What influence variance in any part of the system
will have on the remainder can only be
determined by playing out the final exhaustive and quantitative model.
In retrospect, it seems that the perspective upon -which this model depends is the
insight that the factors determining growth
in the hydrozoa are quite different from
those determining form. The idea that
form is a consequence of the location of
proliferative regions is an attractive one
for many reasons and one slow to die. Although data suggesting other bases of morphogenesis were produced simultaneously
in a number of laboratories, I believe that
Clarkson and Wolpert (1967) were the
first to make explicit the general implications for hydroid morphogenesis of the
dependence of bud genesis on the initiation
of proliferative regions in hydra. They, of
course, concurred with a number of other
investigators in negating this dependence.
The repertoire of cellular alternatives
which account for morphogenesis and
morphostasis in Podocoryne is extremely
limited: cells can be epitheliocyte or amoebocyte; they can remain in place or they
can be motile; they can make or dissolve
perisarc. Certainly, there is a broad spectrum of synthetic activities of which hydroid cells are capable, but these are not,
for the larger part, activities which directly
influence colony form.
Not all form-generating activities have
been considered in the model. The hydranth has been taken as a given, although
I have attempted to show how cell proclivities can account for the steady state
of the polyp. The origin of sexual persons
has been ignored. The model is by no
means exhaustive. My intent has been to
demonstrate that colony morphogenesis and
morphostasis are the consequence of a
small number of variables in cell proclivities, that the building blocks out of which
colony form is constructed are few.
To contend that the basis of morphogenesis is cellular seems to be a truism.
It is, of course, true that animal architecture is constructed of cellular units. It
697
is also a truism to add that any theory
which purports to account for animal form
must eventually be relevant to cellular
characteristics. That the alternative expressions of which cells that influence shape are
capable are few is not, however, a truism.
I corroborate, in this study, the conclusions of Gustafson and Wolpert (1963)
who identified many of the same cellular
proclivities, adhesion or the lack of it,
tendency to form pseudopods, as responsible for the morphogenesis of the sea
urchin pluteus. It begins to appear that
the complexity of animal form, in its
phylogenetic diversity, is structured of a
small number of cellular proclivities reflecting primitive cellular characteristics.
The biochemical variety underlying these
alternatives of cellular behavior may be
diverse, but the repertoire of cell states
which influence form is highly limited.
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