J. Embryol. exp. Morph. 95, 247-260 (1986)
Printed in Great Britain © The Company of Biologists Limited 1986
Regeneration from half lower arms in the axolotl
PETER WIGMORE
Department of Anatomy, King's College London, Strand, London WC2R 2LS, UK
SUMMARY
A technique involving grafting of pieces of skin from the head onto the limb in order to isolate
halves of the limb is described. This technique was used to isolate posterior, anterior, dorsal and
ventral halves of the lower arm. All halves produced regenerates but no part of the limb was able
to produce a high proportion of regenerates with a complete pattern of skeletal structures.
Posterior half stumps regenerated limbs with a mean digit number of 2-7 and had a normal
dorsoventral muscle pattern. Anterior half stumps produced a high proportion of single-digit
regenerates and had a mean digit number of 1-3. Dorsal and ventral half stumps regenerated
limbs with a mean digit number of 2-8 and 2-3 respectively. Hypomorphic regenerates from
dorsal and ventral half stumps often had only dorsal or ventral muscle. These results are in
contrast to those from the upper arm (Wigmore & Holder, 1985) where a complete skeletal and
muscular pattern regenerated from posterior and dorsal halves and hypomorphic regenerates
were obtained from anterior and ventral half limbs.
INTRODUCTION
Limb regeneration in amphibians provides a system for the study of the
formation of a complex pattern of skeletal and soft tissue elements. Information
for this pattern is believed to lie in the cells close to the amputation plane and to be
present in both the dermis and underlying muscles (see review by Tank & Holder,
1981). Several techniques for assaying regenerative ability have shown that this
information is not evenly distributed within the upper arm (symmetrical limbs:
Stocum, 1978; Tank & Holder, 1978; Holder, Tank & Bryant, 1980; irradiation:
Maden, 1979; isolation of half limbs using head skin: Wigmore & Holder, 1985).
These techniques have shown that normal 4-digit limbs can only form if the stump
contains posterior or dorsal tissues. Stumps lacking these tissues produce hypo­
morphic regenerates, anterior halves in particular only regenerate single digits.
The dorsoventral axis, assayed using the pattern of muscles, appears to be
controlled independently of the anteroposterior axis, allowing the formation of
limbs with a normal skeletal pattern but abnormal muscle distribution. The
dorsoventral axis also tolerates the presence of discontinuities in the pattern while
discontinuities present in the anteroposterior axis of the stump result in the
formation of additional structures in the regenerate (Maden, 1980; Holder &
Weekes, 1984,1985; Burton, Holder & Jesani, 1986).
The lower arm has been less-intensively studied but there is some evidence that
the amount of structure regenerating from different halves is different from the
Key words: axolotl, limb, regeneration, pattern formation, muscle.
247
248
P. WIGMORE
upper arm. Surgical removal of the posterior half of the lower arm of the newt
(Goss, 1957A; Dinsmore, 1983) produced regenerates with between 1 and 4 digits
but with a mean digit number of 2-4. Irradiation of half the limb (Goss, 19576;
Maden, 1979) showed that both posterior and anterior halves could produce on
average slightly more than 2 digits but that posterior halves tended to produce
more than anterior halves. The implantation of unirradiated half limbs into
irradiated stumps (Stinson, 1964) or the construction of symmetrical lower legs
(Stocum, 1978) produced a greater difference between anterior and posterior
halves; the posterior half producing approximately three quarters of the normal
limb and the anterior one quarter. Construction of symmetrical lower arms in the
axolotl (Krasner & Bryant, 1980) produced regenerates with on average less than 2
digits from either double anterior or double posterior stumps while construction of
limbs with symmetrical skin produced double posterior regenerates from double
posterior stumps and normal or hypomorphic regenerates from stumps with
double anterior skin (Slack, 1980). These results indicate that in contrast to the
upper arm no part of the lower arm is capable of producing a complete re­
generated limb. Despite the apparent inability of lower arm regenerates to
compensate for deficiencies in the stump this region of the limb shows strong
powers of regulation by the production of supernumeraries when anterior and
posterior tissues are opposed (Tank, 1978). No study of the dorsoventral axis has
been done after manipulations on the lower arm and part of this study was
designed to ascertain whether this axis behaves in the same way in the lower arm as
in the upper.
The present work is the application of a new technique which has been able to
demonstrate differences in regenerative ability in different regions of the upper
arm (Wigmore & Holder, 1985). This technique uses the fact that head skin
contains no positional information affecting limb regeneration (Wigmore &
Holder, 1985) and rapidly heals to a wound made in the limb. These properties are
. used to isolate halves of the arm by grafting head skin onto the medial surface of
amputated stumps from which half of the limb has been removed. The position of
the head skin prevents any contribution from proximal parts of the limb corres­
ponding to the half of the stump that has been removed and restricts regeneration
to that produced by the isolated half. The controls consisted of an identical
operation but without any grafting of head skin. This allows all parts of the limb to
interact around the amputation plane and contribute to the regenerate.
MATERIALS AND METHODS
All experiments were carried out using axolotls (Ambystoma mexicanum), 15-20 cm in length
spawned in the colony at King's College. Animals were kept in tap water in individual containers
and fed on raw heart.
Experimental procedure
During surgery animals were anaesthetized in MS222 (Sigma), after recovery they were kept
at 10°C in the dark for two days in order to facilitate healing. All operations were performed on
Regeneration from half lower arms in the axolotl
249
Fig. 1. Victoria-blue-stained whole mount of a regenerate from a control limb with a
normal skeletal pattern. The digits are numbered from 1 to 4 in an anterior to posterior
direction. The bones are labelled as follows, r, radius; u, ulna; i, intermedium; ra,
radiale; p, prepollicis; b 1/2, fused basal carpals one and two; b 3, basal carpal three;
b 4, basal carpal four; c, centrum; ul, ulnare. Scale bar, 0-25 cm.
the zeugopodium with the arm at right angles to the body and the elbowflexedusing the axes of
the body to determine the position of different halves of the limb (see Burton et al. 1986).
Experimental operations comprised the removal of half the lower arm (either the dorsal,
ventral, anterior or posterior half). Posterior and anterior half stumps included either the ulna or
radius respectively while both forearm bones were included in both dorsal and ventral halves.
The medial wound surface formed by the removal of half the forearm was covered with a
3-4 mm wide strip of skin removed from the forehead which was sutured into place (see
Wigmore & Holder, 1985). The limb was then amputated through the graft and trimmed so that
the amputation plane passed through the middle third of the forearm. Thus the stumps consisted
of either a single forearm bone in the case of anterior and posterior halves or both radius and
ulna in dorsal and ventral half stumps together with approximately 50 % of the circumference of
limb skin and soft tissues, the remainder of the circumference being covered by head skin.
Controls consisted of carrying out the same procedure as described above but without grafting
any head skin onto the limb.
All experimental animals were checked for retention of the graft every few days for the first
few weeks after the operation. Operated animals were left for 60 days after which time the limbs
were amputated, fixed in Bouin's solution, dehydrated and stained with Victoria Blue (Bryant &
Iten, 1974) to show the pattern of cartilages. Fig. 1 shows a control regenerate with a normal
skeletal pattern. Camera-lucida drawings were made of all regenerates. Limbs were then wax
embedded and serial transverse sections were cut at 10 jum and stained with haematoxylin and
eosin. These sections were used to check the position of the graft and to analyse the dorsoventral
muscle pattern. Limbs where the graft was not in the expected location were discarded.
The muscle pattern was examined at two levels, the proximal metacarpals and the midforearm. In normal limbs at the level of the proximal metacarpals, ventral muscle is continuous
across all digits while dorsal muscle occurs as discrete crescents over each metacarpal and a large
vascular sinus is present only on the ventral side (Maden, 1980,1982; Maden & Mustafa, 1982;
Burton et al. 1986). At the midforearm level, the dorsoventral axis can be determined from the
overall muscle pattern and the presence of the pronator quadratus (pq) muscle which runs from
a spine on the ventral side of the ulna towards the radius (Holder & Weekes, 1984; Burton et al.
1986). Head-skin-derived tissue was normally easily recognizable as a fold of tissue on the side of
the limb overlying an extensive region of loose connective tissue. Serial sections of limbs showed
250
P. W I G M O R E
this fold to be continuous with the original head-skin graft. The anatomical nomenclature used
was obtained from Francis (1934) and Grim & Carlson (1974).
RESULTS
Posterior half limbs
Twenty posterior half stumps were constructed all of which produced a
regenerate (see Table 1). Regenerates had between 1 and 4 digits, the most
frequent type (40 % of limbs) having 2 digits. Of the 4-digit limbs all except for one
was normal; the exception was a limb that had a proximally incomplete radius and
lacked phalanges from digits 1 and 2. One 3-digit limb possessed the distal part of
the radius (Fig. 2A) while the remaining 3-digit limbs had only an ulna (Fig. 2B).
All 3-digit limbs appeared to consist of digits 2, 3 and 4. These limbs were
asymmetrical and although lacking anterior structures, had apparently normal
posterior halves. All 2-digit limbs appeared to be digits 3 and 4 (Fig. 2C) but
unlike the 3-digit regenerates six out of eight had a complete radius although this
did not always articulate with the carpals (Fig. 2D). Two regenerates were distally
incomplete, lacking phalanges and having only the proximal ends of metacarpals.
Both single-digit regenerates (Fig. 2E) possessed only an ulna and one was distally
incomplete.
Sections of regenerates from posterior half stumps showed a fold of loose
connective tissue that was normally located on the anterior side. This feature was
continuous with the head skin graft and is assumed to be tissue derived from the
graft. The muscle pattern of all limbs was normal at the forearm level, however,
graft-derived tissue was found to lie in a dorsoanterior location in some limbs and
this may have led to the absence of dorsal muscle in the metacarpal region of two
of the 2-digit limbs. One limb of 3 digits possessed a duplicate radius distally and
the graft-derived tissue in this limb was located anteriorly and ventrally. The
remaining limbs had a normal muscle pattern at the metacarpal level.
Anterior half limbs
Thirteen anterior half limbs were made using the contralateral limbs of the
animals used to make posterior half stumps (see Table 1). Anterior half stumps
regenerated considerably less structure than those from posterior halves with a
mean of only 1-3 digits compared to 2-7. The majority of anterior half stumps
produced only a single digit with a single forearm bone, the radius. Of the three
regenerates with more than 1 digit, the 3-digit limb had both radius and ulna but
Table 1. Analysis of the number of digits regenerating from different halves of the limb
Stump
Posterior half
Anterior half
Dorsal half
Ventral half
No. of limbs
produced
20
13
8
10
No. of digits regenerating
4
3
2
1
6
4
1
2
1
8
2
2
5
2
10
3
2
No
regeneration
1
2
Mean no.
of digits
2-7
1-3
2-8
2-3
Regeneration from half lower arms in the axolotl
2A
C
/
B /
^^^»^
D
251
^ '
y
Fig. 2. Victoria-blue-stained whole mounts showing skeletal patterns regenerating
from posterior half stumps. (A) A 3-digit regenerate lacking digit 1 but with the
remainder of the skeletal pattern being approximately normal. (B) A 3-digit regen­
erate lacking digit 1, the radius and the most anterior carpals (the prepollicis and
radiale) together with the anterior parts of the centrale and intermedium. The
posterior digits and carpals appear normal. (C) A 2-digit regenerate lacking digits 1
and 2 together with the anterior and medial carpals. The radius is also missing but digits
3 and 4, the posterior carpals and ulna appear normal. (D) A 2-digit regenerate similar
to that in (C) but possessing a radius which does not articulate with any of the carpals.
(E) A single-digit regenerate with only an ulna although small pieces of cartilage occur
where the radius would be. Scale bars, 0-25 cm.
the anterior digit appeared unconnected to the posterior pair (Fig. 3A). One
2-digit limb had both radius and ulna (Fig. 3B) while the other possessed only
a radius and consisted of digits 1 and 2 (Fig. 3C). All the single-digit regenerates
had only a radius with two or three carpals, a metacarpal and two phalanges.
Occasionally spikes of cartilage were found lying posterior to the radius in the
position expected of the ulna (Fig. 3D).
It was not normally possible to recognize dorsoventral muscle patterns in singledigit limbs but the distribution of muscle was often asymmetric. Both 2-digit limbs
252
P. W I G M O R E
3Ä"
Fig. 3. Victoria-blue-stained whole mounts showing the skeletal patterns regenerating
from anterior half stumps. (A) A 3-digit regenerate possibly consisting of digits 1, 2
and 3 and also having both radius and ulna. (B) A 2-digit regenerate with both radius
and ulna. (C) A 2-digit regenerate with only a radius. The digits are recognizable as 1
and 2, and the anterior carpals are present (prepollicis, radiale, fused basal carpals one
and two and the anterior part of the centrum). (D) A single-digit regenerate with only
a radius although a small piece of cartilage lies where the ulna would be expected.
Scale bars, 0-25 cm.
Regeneration from half lower arms in the axolotl
253
had dorsal and ventral muscles but the pattern in the 3-digit limb was un­
recognizable. All limbs had graft-derived tissue present on their posterior side.
Dorsal half limbs
Eight dorsal half stumps were constructed and all but one produced a regenerate
(see Table 1), consisting of between 2 and 4 digits with a mean digit number of 2-8.
All regenerates had both a radius and ulna. One 4-digit regenerate was normal
while the remaining two lacked posterior carpals and in one case the phalanges of
digit 4. The 2- and 3-digit limbs appeared symmetrical about their anteroposterior
axis (Fig. 4A,B) but unlike regenerates from posterior half stumps the digits could
not be identified with any certainty.
Sections of regenerates from dorsal half stumps showed graft-derived tissue on
the ventral side of the limb at proximal levels but this tissue tended to be localized
more anteriorly at more distal positions. All of the 4-digit and one 3-digit limb had
normal muscle patterns at both metacarpal and forearm levels although in one
regenerate the hand was folded to enclose the ventral surface. The three re­
maining 2- and 3-digit limbs were either double dorsal (Figs 6, 7) or had dorsal
muscle but no muscle on the ventral side. This was true both at the metacarpal and
forearm levels. One 3-digit limb had three bones in the distal forearm (Fig. 7).
Ventral half limbs
Ten ventral half stumps were completed and all produced a regenerate varying
between 1 and 4 digits (see Table 1) with 50 % of limbs producing 2 digits. Both 4digits limbs had a normal pattern of bones and all other regenerates had both
radius and ulna. The digits of the 2- and 3-digit limbs were not identifiable
(Fig. 5A,B); digits were often not in the same plane as the rest of the limb and
were sometimes fused at the metacarpal level. Single-digit limbs also had both a
radius and an ulna but although having five or six carpals only a single metacarpal
with two phalanges was present distally. One of the 2-digit limbs had three bones
in the distal forearm.
As with the 4-digit dorsal half stump regenerates, 4-digit regenerates from
ventral stumps had normal muscle patterns at both metacarpal and forearm levels.
The remaining limbs had either ventral muscle on both sides of the limb (five
limbs) or ventral muscle on one side and an absence of muscle on the dorsal side
(three limbs). Double ventral limbs had continuous muscle between the digits on
both sides of the metacarpals and two pronator quadratus muscles in the forearm
(Figs 8, 9). One limb, which was double ventral in the forearm lacked any muscle
distally.
Control half limbs
Control consisted of ten anterior, ten posterior, five dorsal and five ventral half
limbs. All but one of these limbs regenerated 4 digits, the exception being a 5-digit
limb produced from an anterior half stump. All regenerates possessed both a
254
P. WlGMORE
4A
4B
5A
5B
Fig. 4. Victoria-blue-stained whole mounts showing the skeletal pattern regenerating
from dorsal half stumps. (A) A 3-digit regenerate with both radius and ulna.
(B) A 2-digit regenerate also with both radius and ulna. Scale bars, 0-25 cm.
Fig. 5. Victoria-blue-stained whole mounts showing the skeletal pattern regenerating
from ventral half stumps. 2-digit regenerates with both radius and ulna. Scale bars,
0-25 cm.
Regeneration from half lower arms in the axolotl
255
radius and an ulna. Some anterior half stumps were seen to have medium bud
blastemas with two apices and this type of stump had the highest proportion of
abnormalities in the final regenerate. The commonest abnormality that was seen
on three posterior half and six anterior half controls was the appearance of an
additional basal carpal between the fused basal carpals one and two and basal
carpal three (Fig. 10). Five anterior limbs and one posterior and one dorsal limb
Fig. 6. Camera-lucida tracing of a section through the proximal metacarpals of a
2-digit regenerate from a dorsal half stump. Dorsal muscle (the extensor digitorum
brevis) is present on both dorsal and ventral sides of the limb, c, metacarpals; shaded
areas, dorsal muscle. The head-skin-derived tissue, marked h, occupies the anterior
side of the hand and its limits are marked by the arrows. x25.
Fig. 7. Camera-lucida tracing of a section through the distal forearm of a regenerate
from a dorsal half stump. Three forearm bones were found at this level and dorsal
muscle is present on both dorsal and ventral sides of the limb. Bones: u, ulna; r, radius;
c, unidentified third bone. Muscles: edc, extensor digitorum communis; ear, extensor
antebrachii radialis; ecr, extensor carpi radialis; h, head-skin-derived tissue, arrows
indicate its limits. x25.
256
P. W I G M O R E
Fig. 8. Camera-lucida tracing of a section through the proximal metacarpals of a
2-digit regenerate from a ventral half stump. Ventral muscle occupies both dorsal and
ventral sides of the limb. Starred areas, ventral muscle; c, metacarpals; h indicates
head-skin-derived material and the arrows its limits. x25.
Fig. 9. Camera-lucida tracing of a section through the distal forearm of a regenerate
from a ventral half stump. Ventral muscle is present on both sides of the limb. Bones:
u, ulna; r, radius. Muscles: feu, flexor carpi ulnaris; pq, pronator quadratus; ps,
palmaris superficialis; f aar, flexor ante brachii; h indicates head-skin-derived tissue and
the arrows its limits. x25.
also had three phalanges on two of their digits. The dorsoventral muscle pattern
was normal at both metacarpal and forearm levels for all control regenerates.
DISCUSSION
This work was carried out to investigate the regenerative potential of different
parts of the lower arm and, in particular, to compare these results with those from
the upper arm (Wigmore & Holder, 1985). Previous authors (see Introduction)
have produced results indicating that the lower part of the limb is not able to
compensate for defects in the stump. This contrasts with the upper arm, where
normal regenerates can be produced from dorsal or posterior half stumps. In the
Regenerationfromhalf lower arms in the axolotl
257
present work all halves of the lower arm produced a high proportion of hypo­
morphic regenerates and the ability of posterior and dorsal tissue to consistently
regulate to produce normal regenerates was not found.
In the upper arm the greatest difference in the amount of skeletal tissue
regenerating was found between anterior and posterior half stumps. The mean
number of digits regenerating from posterior upper arm stumps was 3-8 but only
1*2 regenerated from anterior half stumps. In the lower arm the posterior half still
regenerated more digits than the anterior half but the difference was less marked
(2-7 digits from posterior halves, 1-3 from anterior). Compared with the upper
arm there was little change in the regenerative ability of the lower arm anterior
halves with the commonest regenerate still being a single digit. This was always
associated with a radius although parts of the ulna were occasionally present
(Fig. 3D) and a complete ulna was present in two of the multidigit limbs
(Fig. 3A,B).
Posterior half stumps produced the greatest proportion of normal limbs but the
majority of regenerates (70 %) were hypomorphic due to the absence of anterior
digits and carpals. This contrasts with the upper arm where 85 % of limbs were
normal. The radius was also absent in all 3-digit regenerates but surprisingly was
present in many 2-digit limbs. The posterior parts of hypomorphic limbs from
posterior half stumps appeared normal but the amount of the normal skeletal
pattern present varied, often ending abruptly and sometimes even splitting in­
dividual carpals (Fig. 2B). Similar limbs were made from anterior half stumps
(Fig. 3C) but these consisted of anterior structures and occurred at a much lower
Fig. 10. Victoria-blue-stained whole mount showing the skeletal pattern regenerating
from an anterior half control stump. The digits are numbered from 1 to 4 in an anterior
to posterior direction. Digit 2 is abnormal in having three phalanges, c indicates an
extra basal carpal between basal carpal three and fused basal carpals one and two.
Scale bar, 0-25 cm.
258
P. W I G M O R E
frequency. The two single-digit limbs regenerating from posterior half stumps are
difficult to identify but since they were associated with only an ulna they can
tentatively be identified as digit 4. Single-digit limbs were never produced from
upper arm posterior half stumps. The dorsoventral muscle pattern could be
recognized in multidigit limbs and in these limbs the pattern was normal ir­
respective of whether the limb was from an anterior or posterior half stump.
Regenerates from dorsal and ventral half stumps were similar in their antero­
posterior axis; although a proportion of both types of stump produced normal
limbs, the majority of regenerates, while possessing both radius and ulna, had only
2 or 3 digits. Some digits could be tentatively identified from their articulation with
their carpals, for example in Fig. 4B the digits may be 3 and 4 while Fig. 5A may
show digits 1 and 2. In general however it was not possible to be sure what part of
the limb had regenerated. These limbs therefore differ from the asymmetric
regenerates produced from posterior or anterior half stumps which clearly con­
tained part of the normal anterior-posterior pattern.
The major difference between regenerates from dorsal and ventral half stumps
was in their muscle patterns. Limbs with 4 digits had a normal muscle pattern but
nearly all hypomorphic regenerates had only dorsal or ventral muscle depending
on whether they were from dorsal or ventral half stumps. The muscle present was
either on both sides of the limb making them double dorsal or double ventral or
only occupied one side leaving head-skin-derived tissue on the other (half-ventral
or half-dorsal limbs). Double or half distributions of muscle were present at both
forearm and metacarpal levels. Where double ventral muscle occurred at the
forearm the symmetry of the soft tissues was also reflected in the bones as a
normally ventral spine on the ulna was present on both sides of the bone. In the
upper arm, ventral half stumps regenerated a high proportion of limbs with very
similar muscle patterns to those found here. Upper arm dorsal half stumps
however, nearly always regenerated limbs with a normal dorsoventral muscle
pattern; the exception was one limb that was half dorsal (Wigmore & Holder,
1985). A possible explanation for the normality of muscle patterns from upper arm
dorsal regenerates was that the boundary between dorsal and ventral tissue (see
Meinhardt, 1983) lies dorsal to the humerus in the upper arm. In the experimental
protocol used, this could have led to the inclusion of ventral tissue in dorsal half
upper arm stumps. The higher proportion of regenerates from lower arm dorsal
half stumps with only dorsal muscle present may indicate that this boundary is
located more ventrally in the forearm. Despite the above differences, the pro­
duction of limbs with only dorsal muscle from dorsal half stumps and only ventral
muscle from ventral half stumps in both upper and lower arms is evidence that
these two tissue types act mosaically during regeneration. Results from limbs
symmetrical in the dorsoventral axis (Burton etal. 1986) and the surgical con­
struction of discontinuities in this axis within the stump (Holder & Weekes, 1984)
have provided further evidence for this.
Control limbs had normal dorsoventral muscle patterns for all four types of
stump which was not surprising, as, despite the uneven shape of the stump after
Regeneration from half lower arms in the axolotl
259
removal of half of the limb, there was no barrier to the interaction and incor­
poration of all parts of the stump. The anteroposterior skeletal pattern was also
normal for most posterior, dorsal and ventral half stumps but a frequent abnor­
mality in anterior halves was the presence of an extra basal carpal between basal
carpals two and three (Fig. 10). This bone did not appear in any experimental
regenerates but its occurrence may be linked to the presence of 'double' blastemas
seen on anterior half control stumps. Without the head skin that was present in
experimental stumps, control stumps had two transverse amputation plançs at
slightly different proximodistal levels. It seems likely that both planes start to
produce blastemas which subsequently fuse to form a single regenerate. The
presence of the extra basal carpal maybe evidence that the integration between the
two blastemas was not perfect.
The technique described here enables the isolation of half stumps with head
skin. This tissue, despite contributing cells to the regenerate, appears to have no
morphogenic effects (Wigmore & Holder, 1985) but by preventing any contri­
bution from cells on the side of the limb that was removed we were able to show
differences in the regenerative ability of different parts of the limb. The major
difference between upper and lower arm results was the inability of any part of the
lower arm to produce a high proportion of normal regenerates. In the lower arm
posterior half stumps can produce a variable amount of the anterior-posterior
axis from 1 to 4 digits while the anterior half can produce between 1 and 3 digits
but with a high proportion of single-digit regenerates. The variation in the
completeness of the pattern is possibly due to variations in the position of the
medial cut made in constructing these stumps.
Dorsal and ventral lower arm halves showed approximately equal regenerative
potential. Many of these limbs were similar to regenerates from upper arm
operations in having only dorsal or ventral muscle. It is likely that the dorsoventral axis is specified in a similar way in both the upper and lower parts of the
arm. The ability to regenerate the pattern of the normal limb is thought to be due
to the holding of different positional values by cells in different parts of the limb.
A possible explanation for the difference between upper and lower arms in the re­
generation of the anteroposterior axis suggested by Maden (1979) is that positional
values are clustered in the posterior dorsal quadrant of the upper arm but are more
diffusely distributed in the lower arm. The present results fit this interpretation but
further work is required to define the interactions between different regions of the
limb in making the normal pattern.
I would like to thank Rosie Burton, Dick Glade and Nigel Holder for many useful comments
on this work and Meena Jesani for her expert technical assistance. The project was supported
financially by the Science and Engineering Research Council.
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BURTON,
(Accepted 28 February 1986)