VII.
LIMB MORPHOGENESIS
321
III. Proximo-distal Sequence in Limb Development
There seems to be general agreement about at least one aspect of limb
development, namely the sequence of origin of limb parts. Until 1948
there was no definitive information about the sequence in which such
parts formed. Many descriptions of limb development stated that the
initial bud represented the digital structures and that proximal parts
were added behind these (Milaire, 1956, feels that this may still be the
situation in mammalian limb development). Earlier experiments of
Balinsky (1935) had indicated that this might not be the situation in
Triturus. Mesodermal components of successively older limb buds were
isolated and grown in enucleated orbits of the eyes of older hosts.
Mesoderm from the younger buds formed proximal parts while more
distal parts appeared when progressively older buds were used. Saunders
(1948) described convincing data to support his contention that limb
parts form in a proximo-distal sequence in the chick embryo. He had
become curious about the morphogenetic role of the distinctive ectodermal thickening (the apical ridge which has been discussed so much
above) seen along the distal edge of limb buds. When the ridge was
to test some of these ideas are' now in progress both in my laboratory
and in Bell's.
It is evident from this discussion as well as that which has preceded
it that our knowledge of mechanisms is still rudimentary and still
restricted to a biological level. Attempts to reach a molecular level for
an elucidation of mechanisms have, thus far, been restricted to studies
of the histochemistry of developing limbs. The most striking observation
which has come from histochemical studies has been the high alkaline
phosphatase content of the apical ectodermal ridge in the limb buds of
rodents (Hinrichsen, 1956; McAlpine, 1956; Milaire, 1956). Milaire, who
has deduced an amazing amount from histochemistry (including the
idea of interaction between the ectoderm and mesoderm) interpreted
the high alkaline phosphatase of the ectodermal ridge to be an indication
of inductive activity. Similar intense alkaline phosphatase could not be
detected in the limb buds of chick embryos (Loewenthal, unpublished
work). Presumptive limb muscle cells were characterized (v. Weel, 1948)
by a concentration of ascorbic acid. Histochemical studies have not, to
this date, been particularly revealing. The stage is set, though, for more
sophisticated approaches. As yet there have been no attempts to study
the basement membrane by means of electron microscopy or other
modern methods. Even though there are still some disagreements about
basic biological mechanisms, the time is now ripe for new analytical
approaches.
LIMB MORPHOGENESIS
321
III. Proximo-distal Sequence in Limb Development
There seems to be general agreement about at least one aspect of limb
development, namely the sequence of origin of limb parts. Until 1948
there was no definitive information about the sequence in which such
parts formed. Many descriptions of limb development stated that the
initial bud represented the digital structures and that proximal parts
were added behind these (Milaire, 1956, feels that this may still be the
situation in mammalian limb development). Earlier experiments of
Balinsky (1935) had indicated that this might not be the situation in
Triturus. Mesodermal components of successively older limb buds were
isolated and grown in enucleated orbits of the eyes of older hosts.
Mesoderm from the younger buds formed proximal parts while more
distal parts appeared when progressively older buds were used. Saunders
(1948) described convincing data to support his contention that limb
parts form in a proximo-distal sequence in the chick embryo. He had
become curious about the morphogenetic role of the distinctive ectodermal thickening (the apical ridge which has been discussed so much
above) seen along the distal edge of limb buds. When the ridge was
to test some of these ideas are' now in progress both in my laboratory
and in Bell's.
It is evident from this discussion as well as that which has preceded
it that our knowledge of mechanisms is still rudimentary and still
restricted to a biological level. Attempts to reach a molecular level for
an elucidation of mechanisms have, thus far, been restricted to studies
of the histochemistry of developing limbs. The most striking observation
which has come from histochemical studies has been the high alkaline
phosphatase content of the apical ectodermal ridge in the limb buds of
rodents (Hinrichsen, 1956; McAlpine, 1956; Milaire, 1956). Milaire, who
has deduced an amazing amount from histochemistry (including the
idea of interaction between the ectoderm and mesoderm) interpreted
the high alkaline phosphatase of the ectodermal ridge to be an indication
of inductive activity. Similar intense alkaline phosphatase could not be
detected in the limb buds of chick embryos (Loewenthal, unpublished
work). Presumptive limb muscle cells were characterized (v. Weel, 1948)
by a concentration of ascorbic acid. Histochemical studies have not, to
this date, been particularly revealing. The stage is set, though, for more
sophisticated approaches. As yet there have been no attempts to study
the basement membrane by means of electron microscopy or other
modern methods. Even though there are still some disagreements about
basic biological mechanisms, the time is now ripe for new analytical
approaches.