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SUMMARY
The cuticles of Pandinus imperator and Scorpiops hardwickii have been examined
histologically and histochemically. They consist of a two-layered epicuticle, a hyaline
exocuticle, a quinone-tanned exocuticle, an impregnated but untanned exocuticle,
and on the inner surface an unmodified procuticle. The hyaline exocuticle is, however,
single in Scorpiops and double in Pandinus. With the exception of the hyaline exocuticle
the cuticles therefore conform to the basic arthropod pattern.
The quinone-tanned exocuticle closely corresponds with that of insects but the
hyaline exocuticle seems to have no counterpart in other arthropods. The sulphur
content of the cuticle has been determined and it is probable that sulphur is present
only in the hyaline exocuticle.
The statement by Krishnan that chitin is present in the epicuticle is critically
examined and a possible explanation of the differences which have been reported
between the cuticles of different scorpions is offered.
INTRODUCTION
JOp.
FIG. I . Transverse section of a fully hardened dorsal abdominal sclerite of
Pandinus imperator.
FIG. 2. Transverse section of the cuticle of a fully hardened adult Pandinus imperator, to show
the transition from sclerite to arthrodial membrane.
tOji
A
—.cuticulin
layer
_ — . "
• : - : — " ^
—enttocutfclt
~ ZTZ,
FIG. 3. A, transverse section of the cuticle of a newly born young of Pandinus imperator.
B, greatly magnified view of the region enclosed by the rectangle in A (not to scale).
by groups of pore canals which take up the acid fuchs.in of Mallory (fig. 4).
The pore canals cannot be traced into the endocuticle and it can be only
assumed that they are present and stain the same colour as the endocuticle.
Laminae are present throughout the endocuticle and are regular in appearance.
Dermal gland ducts are present and are more closely spaced than in the adult.
They arise from enormous gland-cells situated between the epidermal cells.
The ducts have the general form of those of the adult cuticle (fig. 1).
The chemical composition of the cuticle. The various histochemical reactions of
the fully hardened cuticle are summarized in table 1. The positive chitosan
reaction (treatment with caustic potash followed by the application of sulphuric
acid and iodine) indicates that the chitin is the main constituent of the cuticle
beneath the hyaline exocuticle. The hyaline exocuticle did not give a positive
reaction.
After treatment with potash all the region reacting with the iodine and sul-
phuric acid dissolved rapidly in 3 % acetic acid; this further indicates the
4° Kennaugh—Cuticles of Scorpions
presence of chitin. The hyaline exocuticle did not dissolve. Sections of
cuticles after treatment with potash tended to split at the margin of the
quinone-tanned and hyaline exocuticle. The hyaline exocuticle does not be-
come reactive towards staining and chemical tests after potash treatment.
apicuticle
.outer region of
hyaline exocuticle
TIT I T I i r r m region of
hyaline exocuticle
_ pore canal
s— endocuticle
•;V.- epidermis
1OM.
TABLE I
All these observations are in agreement with those of Krishnan (1953) for the
cuticle of Palamneus swammerdami.
Positive Millon and xanthoproteic reactions were given by the cuticulin
layer and quinone-tanned exocuticle. The reaction given by the colourless
exocuticle was more feeble and no reaction was detected in the endocuticle.
With Morner's reagent no reaction was given with any part of the cuticle.
This is in agreement with the observations of Krishnan (1953), who did not
find tyrosine to be present in the cuticle of Palamneus. The argentaffin
Kennaugh—Cuticles of Scorpions 47
reaction was strongest in the quinone-tanned exocuticle; it was probably due
to the presence here of quinones. A less strong reaction was given by the
cuticulin layer and only a very pale brown colour was seen in the colourless
exocuticle. The endocuticle did not react.
The staining reactions of the cuticle before and after treatment with
diaphanol are interesting. Before treatment the quinone-tanned exocuticle
is not stained with any of the components of Mallory's triple stain but the
colourless exocuticle and endocuticle stain red and blue respectively. After
10 weeks' treatment with diaphanol the cuticle was bleached and the staining
reactions modified. The bleached tanned exocuticle stained with aniline blue
and both the colourless exocuticle and endocuticle stained light blue. This
recalls the condition seen in the cockroach, Periplaneta americana, where it
was found by Dennell and Malek (1956) that treatment of the adult cuticle or
any hardened cuticle caused it to stain like the untanned cuticle. After more
prolonged treatment with diaphanol the whole region of the cuticle internal
to the hyaline exocuticle, instead of staining a definite pale blue, becomes a
dull blue-gray. This may be related to the fact that the cuticle now consists of
purified chitin. The hyaline exocuticle is not affected in any way by this pro-
longed treatment and it can therefore only be assumed that quinone tanning
is not an important feature of this region of the cuticle. In cold strong nitric
acid most of the cuticle dissolves fairly readily but the epicuticle and hyaline
exocuticle do not dissolve. With hot or boiling concentrated nitric acid the
epicuticle and hyaline exocuticle dissolve slowly. These outer layers do not
dissolve in 6N hydrochloric acid at ioo° C. even if the treatment is prolonged
for 14 days. It was difficult to identify the cuticulin layer after this treatment,
but the paraffin layer is still present and stains with aniline blue. These
observations and similar ones of Krishnan (1953) seem to be consistent with
the view that the stability of the hyaline exocuticle is attributable to sulphur
bonding. Using both the alkaline lead acetate and nitroprusside reagents,
Krishnan (1953) has been able to show the presence of sulphur in the inner
epicuticle (hyaline exocuticle) of Palamneus. In the present work, however,
neither of these tests gave positive results. Pearse (1953) records that the lead
acetate reaction is not given by tissues fixed in either formalin or alcohol.
Pearse also found that the colour given by the nitroprusside reagent on tissues
fixed in formalin or alcohol was invisible in sections. In the light of these
observations the lack of positive results cannot be taken as indicating the
absence of sulphur. Chromatographic analysis of the cuticle does not reveal
cystine or other amino-acids containing sulphur. If the stability of the hyaline
exocuticle is due to sulphur bonding, the apparent absence of sulphur is easily
explicable. Krishnan (1953) found that treatment with alkaline sodium
sulphide, a reagent said to rupture sulphur bonding, caused the inner epi-
cuticle (hyaline exocuticle) to become reactive to stains. This observation
would seem to point to the presence of sulphur, but similar reactions were not
given by the cuticle of Pandinus.
In view of the failure to detect sulphur by histochemical means in the
48 Kennaugh—Cuticles of Scorpions
cuticle of Pandinus, it was decided to carry out a gravimetric analysis of
the cuticle according to the method given by Hawk and others (1954). Pieces
of cuticle were taken from various regions of the body and dried to constant
weight in hot air. The cuticle was then oxidized by the application of con-
centrated nitric acid and hydrogen peroxide, to which was later added
bromine. This treatment converts the sulphur to sulphate, which is estimated
as the barium salt. A control experiment was carried out and the difference in
sulphate determined. By this method it was found that the total sulphur con-
tent of the cuticle was about 4"6%. This appears to be the only information
available on the sulphur content of scorpion cuticles. Lafon (1943) records
that in Limulus the sulphur content is 2-85%. By comparison with Pandinus
the sulphur-containing region of the cuticle of Limulus is much thinner than
that of Pandinus and is therefore possible that this alone may account for the
difference between these figures.
Intense colouring with Sudan black B is confined to the paraffin layer of
the epicuticle, which shows as a separate black layer. The inner region of the
hyaline exocuticle is coloured to a certain extent as is the quinone-tanned
exocuticle. The colourless exocuticle and endocuticle take up very little colour
and it is doubtful whether this is significant in these two layers.
The results recorded in this section indicate that except for the presence of
a hyaline exocuticle, which presumably contains sulphur, the cuticle of
Pandinus bears a strong resemblance to the cuticle of insects. It contains
chitin and protein, and a quinone-tanned exocuticle is present. The thin sur-
face layer of the epicuticle, which colours strongly with Sudan black B,
appears to correspond to the paraffin layer of the insect cuticle.
REFERENCES
DENNELL, R., and MALEK, S. R. A., 1955. Proc. Roy. Soc, 143, 427.
1956. Ibid., 144, 545.
HAWK, P. B., OSER, B. L., and SUMMEHSON, W. H., 1954. Practical physiological chemistry.
13th ed., London (Churchill).
KRISHNAN, G., 1953. Quart. J. micr. Sci., 94, 11.
1954. Ibid., 95, 371.
, RAMACHANDRAN, G. N., and SANTAMAN, M. S., 195s- Nature, 176, 557.
LAFON, M., 1943. Bull. Inst. Oceanogr., No. 850.
PEARSE, A. G., 1953. Histochemistry, theoretical and applied. London (Churchill).
RICHARDS, A. G., 1951. The integument of arthropods. Minnesota (University Press).
SHRIVASTAVA, S. C , 1954. Current Science, 23, 363.
1955. Ibid., 24, 24.