J. Exp. Biol. (1965), 43, 385-395
With 1 plate and 4 text-figures
Printed in Great Britain
385
THE SITE AND PERMEABILITY OF THE FILTRATION
LOCUS IN THE CRAYFISH ANTENNAL GLAND*
BY LEONARD B. KIRSCHNER AND STANLEY WAGNER
Department of Zoology, Washington State University, Pullman, Washington
(Received 19 February 1965)
INTRODUCTION
Study of the physiology of excretory organs in the invertebrates has lagged far
behind that in vertebrates, and questions for which answers have long existed for the
latter are still moot for most forms. Recent research on the crayfish antennal gland
has shown that this organ operates on the same general plan as the vertebrate kidney;
a primary ultrafiltrate formed somewhere in the organ is modified by reabsorption,
and possibly by secretion, to form a final urine (Riegel & Kirschner, i960). Thus
inulin is excreted by the crayfish, the urine concentration being usually higher than
that in blood. By analogy with the vertebrate kidney the concentration of this compound may be attributed to the reabsorption of water somewhere in the tubular
lumina. Glucose is not normally excreted, but appears in the urine when blood
concentrations exceed about 200 mg. per cent. Animals treated with phloridzin also
excrete glucose, and the maximum urine to blood concentration ratio (U/B) is about
the same as for inulin. It also appears that chloride reabsorption (Peters, 1935;
Riegel, 1963) and dilution of the urine (Riegel, 1963) occur in the tubular portion of
the antennal gland. Some evidence also points to the urinary bladder as a site of
dilution by solute reabsorption (Kamemoto, Keister & Spalding, 1962). Each of these
phenomena has also been described in the kidneys of fresh-water vertebrates (e.g.
Smith, 1950), suggesting the analogy in function mentioned above.
Little is known, however, about the details of antennal-gland physiology. For
example, the filtration locus has not been demonstrated, although the coelosomac is
sometimes cited because, like Bowman's capsule in the vertebrate nephron, it is the
most proximal portion of the organ. And we have no data on the ' porosity' of the
filtration site; that is, on the molecular size compatible with free filtration. The sites
and mechanisms by which most of the filtered material is reabsorbed are unknown,
and we are ignorant of what compounds may be added by secretion from blood to
tubular urine.
As a result of intensive studies on the vertebrate kidney during the past 30 years we
possess techniques for approaching many of these problems. The availability of
radioactive inulin may prove to be particularly useful. There exists a body of evidence
that the behaviour of this compound is similar in both the antennal gland and the
vertebrate kidney; that is, that it is filtered into the tubular lumen, is excluded from
• This investigation was supported by funds for medical and biological research, State of Washington
Initiative Measure 171, and by a grant (G12471) from the National Science Foundation.
25
Exp. Biol. 43, 2
386
LEONARD B. KIRSCHNER AND STANLEY WAGNER
the cells, and is neither secreted nor reabsorbed after filtration. Thus, in several
species of crayfish:
1. Inulin is excreted in the urine, and the U/B is usually 2-3 (Riegel & Kirschner,
i960). The U/B for glucose in phloridzinized animals is also about 2-3, indicating that
this value may be characteristic of a compound that is freely filtered but not reabsorbed.
2. The U/B is essentially constant when the inulin concentration of the blood is
varied over three orders of magnitude (Riegel & Kirschner, i960).
3. Micropuncture studies show that the inulin concentration of the tubular fluid
in the coelomosac and labyrinth is nearly the same as that in blood. This situation
must obtain if free filtration occurs in one of these regions (Riegel, unpublished
experiments).
Such observations indicate that inulin may be used as a reference compound in
studies on the antennal gland as in the vertebrate kidney. The behaviour of other
compounds in various regions of the organ may be assessed by comparing their
concentrations with the concentrations of inulin either in the tubular fluid taken by
micropuncture or in extracts of entire regions. Thus if a test-compound is injected
into the blood the ratio of its blood concentration to that of inulin ought to remain
unchanged in any portion of the antennal gland that treats both compounds in the
same fashion. If the ratio of the test-compound to inulin is not the same as in the
blood the behaviour of the two compounds obviously differs in this region, and the
nature of the difference may provide information about the organ's role in handling the
test-compound.
The observations described in this paper are concerned exclusively with questions
concerning the nitration site. They suggest that the primary ultrafiltrate is, in fact,
formed in the coelomosac, and provide an estimate of the permeability of the filtration
locus.
METHODS
Most of the animals were specimens of Pacifastacus sp. collected locally; a few
experiments were conducted on Orconectes virilus obtained from a commercial
supplier. They were stored at 150 C. in aerated tap-water and were fed twice weekly.
During the period of storage the animals lived in a large porcelain holding tank and
were viable for at least 5 months under these conditions. Individuals isolated for
experiments were placed in small plastic boxes containing about 500 ml. tap-water.
Test-compounds were injected into the sternal sinus, and blood and urine samples
were taken as described in Riegel & Kirschner (i960).
In many of the experiments described the antennal glands were dissected to provide
pieces of coelomosac, labyrinth and tubule. The animals were packed in chipped ice
to immobilize them, the antennal glands were quickly removed and samples of tissue
were taken from the three regions. Both glands could be dissected under a binocular
microscope in 5-7 min. Contrast between coelomosac and tubule (which are physiologically separated by the labyrinth, but morphologically contiguous) was enhanced
by prior injection of Evans's blue (ioo/jg./gm. animal) since the dye becomes concentrated in the coelomosac (cf. Results). The labyrinth was easily distinguished by
its natural green colour. Most tissues were homogenized in and deproteinized by
Somogyi's reagent (Somogyi, 1930), but those prepared for experiments with serum
albumin were treated differently, as described below.
Filtration locus in crayfish antemtal gland
387
Radiochemical procedures
One group of experiments involved a study of the pattern of excretion of polymers
of different sizes and polarity. The compounds used were inulin-carboxy-^C, two
dextrans labelled with M C (one with a molecular weight range of 15,000-20,000, the
other 60,000-90,000), and human serum albumin labelled with 126I. In these experiments blood and urine samples were taken, and aliquots of known volume were
plated on aluminium planchets with no further treatment. After drying they were
counted with a gas flow GM tube through an ultra-thin window. The very low energy
X-ray emitted by m I was found to be detected more efficiently by the GM tube than
by a scintillation crystal.
Some experiments involved the use of pairs of isotopes. For those in which 14C and
3
H were used aliquots of the deproteinized extracts were pipetted into Bray's solution
(Bray, i960) and counted in a two-channel, liquid scintillation counter. Standards
were prepared in the same way for every experiment, hence there was no need to
apply quenching corrections. Tissues containing m I-labelled protein and 14C-labelled
carbohydrate were treated as follows. One aliquot of an aqueous extract of tissue
(blood, antennal gland or urine) was plated on an aluminium planchet, dried, and the
total isotope (12SI + 14C) was counted with the thin-window GM tube. A second sample
was deproteinized as described above and centrifuged to remove the protein. An
aliquot of the supernatant fluid was plated, dried, and counted with the GM tube.
Nearly all of the remaining counts were due to 14C dextran, but a small correction was
applied for m I remaining in the supernatant. This quantity was determined in a
separate experiment by dissolving the albumin and measuring the fraction remaining in
solution after precipitation. In ten trials the range was 0-7-2-6% of the total. The mean
value (1 -4 ± 0-5 %) was used as the correction factor. It was appreciable only in extracts
of the coelomosac.
Microscopic observations
Proteins conjugated with dyes were observed microscopically in order to delimit
their distribution in the antennal gland. Animals were injected with Evans's blue
(which becomes bound to blood protein), or with goat globulin conjugated with
fluorescein. Both antennal glands were excised between 2 and 24 hr. later and
immediately plunged into isopentane cooled to — 160° C. with liquid nitrogen. In the
early experiments, including all those with Evans's blue, the organs were embedded
in paraffin in vacuo, and the paraffin sections were mounted conventionally, i.e. by
floating on warm water. This resulted in loss of most of the dye (see section on
Results). Dry mounting (Branton & Jacobson, 1962) produced too much tissue
distortion in our hands, but the following technique was found to give excellent results.
The frozen organs were either dried as above or were transferred to vials of anhydrous
acetone at — 500 C. and subjected to desiccation by freeze-substitution (Pearse, 1961)
for 6-9 days. The tissues were then fixed by transferring them, still in acetone, to
4° C. overnight. They were then embedded in paraffin (m.p. 52-55° C). Sections
were mounted on slides by the method suggested by J. F. Danielli (Harris, Sloane &
King, 1950). After sectioning, ribbons were placed on warm (40° C.) mercury until
they flattened. An albumin-coated slide was then placed over the sections and permitted
to float there for about a minute. The slide was then lifted from the mercury with
25-2
388
LEONARD B. KIRSCHNER AND STANLEY WAGNER
flattened sections adhering. After deparaffining, a drop of fluorospar was added before
placing the coverslip on the slide. Sections were studied with a Leitz Ortholux
microscope fitted for either phase-contrast or ultraviolet optics.
RESULTS
Excretion of polymers
To attempt to estimate the permeability of the filtration site a series of experiments
was run with a group of polymers of increasing size. Text-fig, i shows the time-course
for excretion of inulin (M.W. 5000) as a function of time. The behaviour of dextran
of low molecular weight (15,000-20,000) (LMWD), shown in Text-fig. 2, is very
48 50 24
-
O
40 -
5
s
o
30
I
i
1
2
4
6
X-
§
c
20
~ ~~x
X—
10
-
~^
l
12
24
36
48
x
I
1
i
60
72
84
Hours
Text-fig. 1. Concentration of inulin in blood (— x —) and urine (— x —) following injection
into the ventral haemocoel at o hr. The inset shows the time-course during the first 6 hr. The
U/B varied between z-i and 2'5 during the period 24—72 hr.
similar. In a series of ten determinations on three animals the mean U/B for this
dextran was 3-0+1-5 (s.D.). In a similar experiment using inulin the mean U/B for
twenty-three experimental periods in ten animals was 2-2 ± 1-2 (s.D.). The difference
between mean values for LMWD and inulin is not significant (o-i > P > 0-05), and
hence the compounds appear to behave similarly.
Text-fig. 3 shows the excretion pattern for a larger dextran (HMWD) with a
molecular weight range of 60,000-90,000. This compound too is excreted, and as with
the LMWD the concentration in the urine exceeds that in the blood. Although the
general pattern of excretion resembles that of the other two, the U/B appears to be
somewhat lower. In a series of sixteen determinations made on three animals the
mean U/B was 1-35 + 0-70 (s.D.). The difference between this value and the mean U/B
Filtration locus in crayfish antennal gland
389
for LMWD or inulin is statistically significant (P < o-oi), although the ranges of
values overlap.
These data suggest that filtration of compounds as large as HMWD may be
restricted, but the use of small groups of animals made it desirable to examine the
question by another method. Four animals were injected with a mixture of HMWD-14C
and inulin-3H. The animals were run in pairs at different times; two were O. virilus,
the other two Pacifastacus sp. Blood and urine samples were removed as nearly
simultaneously as possible and assayed for the isotopes. Another three animals (two
O. virilus and one Pacifastacus sp.) were injected with LMWD-14C and inulin-3H. The
125
100 -
eo
75
8
I"
1
1
I
&
50
—
|
I
1
1
48
•
—
1
,
I
72
Hours
Text-fig. 2. Concentration of low molecular weight dextran in blood (— •—) and urine
(— • — ) following injection of 052 mg. into the ventral haemocoel. The U/B varied between
2-8 and 6-o during the period 24-72 hr.
results are shown in Table 1. For each group the mean ratio of 14C/3H (i.e. dextran/
inulin) in blood is shown in column 4. If both compounds are treated identically this
ratio should be unchanged in the urine. The data in columns 5 and 6 show that this is
nearly true for LMWD-injected animals, but not for the others which excreted the
HMWD only about 69% as fast as inulin.
Excretion of protein was even more restricted. Text-fig. 4 shows the urine and blood
concentrations of human serum albumin-126! following its injection into the haemocoel.
It can be seen that the concentration in the urine was always well below that in blood.
Three animals were injected with labelled albumin, and the mean U/B in seven
measurements was 0-50 ± 0-18 (s.D.). In no case was the concentration in the urine as
great as that in blood. Goat serum globulin (MW about 170,000) labelled with
fluorescein was injected into four animals and no fluorescence appeared in the urine.
In addition, Evans's blue was injected into a series of animals but was not excreted
39°
LEONARD B. KIRSCHNER AND STANLEY WAGNER
in the urine even when the blood concentration was so high that a dilution of 20-fold
was necessary to obtain a spectrophotometric reading. This dye is known to be bound
to plasma protein in vertebrates, and appears to be bound in crayfish blood since
deproteinization left none in the supernatant fluid (unpublished observations).
1200 -
1000 -
Text-fig. 3. Concentration of high molecular weight dextran in blood (—O—) and urine
( — O — ) following injection of 3-0 mg. into the ventral haemocoel. The U/B varied between
0-7 and 1-7 during this experiment. The first urine sample was taken early and the low concentration may have been the result of dilution by bladder urine formed before the dextran was
injected.
Table 1. Blood and urine concentration of dextran and inulin
(See text for description of the experimental protocol. The U/B ratio (column 6) is a ratio of
dextran/inulin values, i.e. column 5/column 4.)
Dextran
HMWD
LMWD
No. of
animals
4
3
No. of
sample
periods
Dextran : Inulin ratios
Blood
Urine
U/B±S.D.
204
o-6o±o-i9
0-92 ±0-24
16
7
Concentration of polymers in the coelomosac
Although Evans's blue was not excreted in the urine it appeared in the antennal
gland as shown in PI. 1,fig.1. In this experiment the gland on the left was taken from
an untreated animal. The coelomosac shows as a darker region near the centre of the
dorsal surface. It is light brown, and stands out from the surrounding tubule, which
is white. The outer rim of the organ, shown in the photograph as a very light border,
Filtration locus in crayfish antennal gland
391
comprises the labyrinth, which is distinctly green. The gland on the right was taken
from another animal which had been injected with 35 fi\. of a 0-5% aqueous solution
of Evans's blue. The organ was removed 24 hr. after injection and immediately
immersed in a physiological saline. The dye was perceptible over the entire surface,
but was obviously very concentrated in the coelomosac. This suggested either that the
coelomosac is more vascular than other regions, or that the dye concentration was
100 -
.5
50 -
.a
<
Fext-fig. 4. Concentration of albumin in blood (—O—) and urine ( O — ) following
injection of 0-024 m g - into t n e ventral haemocoel. T h e U/B was 0 5 2 at 24 hr. It was o-6i
after 98 hr. when the blood concentration had fallen to 1 8 4 /ig./ml.
Table 2. Albumin-inuKn ratios in blood and antennal gland
(See text for experimental protocol. Values for the ratios have been normalized to a blood
value of i-oo in order to facilitate comparison of the two animals.)
Albumin : Inulin ratios
Animal
Time
hr.
2-5
6o
Gland
Right!
Left J
Right)
Left J
Blood
Coelomosac
Labyrinth
W' o
°53
Tubule
0-58
o-57
(17- 7
I 12-3
o-6o
o-57
1-29
(47- 3
o-45
higher in this compartment than elsewhere. Since Evans's blue is bound to protein
the second alternative would indicate that concentration of protein had occurred. To
test this possibility two animals were injected with a mixture of uC-labelled inulin
and m I-labelled human serum albumin. Each animal received o-io ml. of a solution
containing 2-2 fiC 14C and 13-1 fiC. m I . One animal was sacrificed z\ hr. later, the
other 6 hr. In both cases blood samples were removed just before the antennal glands
were removed. The ratio of albumin to inulin was determined in the blood, and in the
coelomosac, labyrinth and tubules for each organ. Values for these ratios appear in
Table 2, and it is obvious that the albumin was much more concentrated in the
coelomosac than in the blood or in other parts of the organ.
392
LEONARD B. KIRSCHNER AND STANLEY WAGNER
If the concentration of protein in the coelomosac is related to filtration, visualization
of the protein might provide evidence concerning the location of the filtration site.
Cross-sections of organs taken from Evans's blue-injected animals showed that the
dye was concentrated exclusively around the tubular lumina in the coelomosac.
However, the colour was not sufficiently intense to warrant photographing, probably
due to dye loss when sections were floated on water (see Methods). Instead, goat
serum globulin conjugated with fluorescein was injected into the haemocoel of a
series of animals and sections of antennal gland were studied with an ultraviolet
microscope. Plate i, figs. 2 and 3, shows a pair of photomicrographs from a single
experimental animal. The latter had been injected with o-i ml. of globulin solution
and the glands were removed 2 hr. later. The labyrinth and tubular portion show only
punctile spots of fluorescence, possibly indicating the location of blood vessels. In
contrast, fluorescence was intense in the coelomosac. The protein is obviously concentrated around the tubular spaces and nowhere else. It is especially noteworthy
that the tubular lumina are completely devoid of fluorescence. On the other hand, the
peritubular cells appear to be completely saturated with the protein.
DISCUSSION
The data presented here indicate that the filtration site in the crayfish antennal
gland is freely permeable to compounds with molecular weights less than 20,000,
while the restriction on excretion of HMWD suggests that restraint on filtration begins
in the molecular weight range 50,000-100,000. Wallenius (1954) found that the
limiting molecular weight for detection in the urine of several mammalian species
varied between 37,800 and 62,700. Some restraint on filtration could be demonstrated
at any molecular weight greater than 5000. Thus, evidence at both ends of the range
of molecular sizes indicates that the permeability of the antennal gland exceeds that
of the vertebrate glomerulus. However, comparison of the excretion patterns for
serum albumin and for HMWD shows that molecular size cannot be the sole parameter governing polymer behaviour. The two compounds have approximately the
same molecular weight, yet the carbohydrate is excreted three times as rapidly as
albumin. If permeability of the filter is involved then factors such as molecular
structure or charge must account for this difference. However, it is also possible that
protein is filtered, then completely reabsorbed at low blood concentrations as has been
demonstrated for haemoglobin in vertebrates (Bayliss, Kerridge & Russell, 1933;
Lippman, Ureen & Oliver, 1951). Since we have never seen either Evans's blue or
fluorescence in any tubules after injection into crayfish the first alternative seems the
more likely. Whatever the mechanism underlying this selectivity its significance is
obvious. The data suggest an approximate upper MW limit for excretion of protein of
150,000. Like the dextran measurements this shows that the antennal gland is
appreciably more permeable than the glomerulus. However, nearly all the protein in
crayfish blood is haemocyanin with a reported MW of about 875,000 (Goodwin, i960),
so these estimates of ' porosity' are commensurate with conservation of blood protein.
Our data also support suggestions that the coelomosac is the site of formation of a
primary ultrafiltrate. The observation that LMWD is handled similarly to inulin in
this region is in accord with this observation, as is Riegel's demonstration that the
Filtration locus in crayfish antennal gland
393
coelomosac tubular fluid contains inulin in virtually the same concentration as blood.
Moreover, the fact that the protein circulating in the blood becomes highly concentrated
in the coelomosac is compatible with filtration at this site. Indeed, it is difficult to
rationalize this observation on any other basis.
The exact site of filtration cannot be delimited from these studies. Concentration
of protein, as shown in PI. 1, fig. 3, clearly occurs only in the peritubular regions. The
fluorescent dye appears to be dispersed throughout the peritubular cells rather than
being confined to the vascular spaces bathing them. This would create some interesting
problems regarding the mechanism by which a filtrate is formed. If the filtrate is, in
fact, formed across the luminal border of the peritubular cells the mechanism must
differ considerably from that in the vertebrates. It is worth noting that such a site for
filtrate formation differs from that suggested by Kummel's electron microscopic
observations (1964), but it might provide a rationale for the appearance of apical
vacuoles in these cells during urine formation (Peters, 1935).
Concentration of protein in the coelomosac raises another question. Our albumin/
inulin ratios appear to suggest that 90-95% of the water entering the coelomosac as
blood must be filtered and pass into more distal regions of the organ.
If the colloid osmotic pressure of crayfish blood reported by Picken, (about
10 mm. Hg) were all due to protein, filtration of this magnitude would be impossible.
However, calculations based on published values show that blood protein cannot
contribute more than a fraction of this value. Thus a reasonable mean protein concentration is about 4% (Florkin, i960), most of it haemocyanin with a MW of about
875,000 (Goodwin, i960). On this basis the blood protein itself contributes less than
1 mm. Hg. The much larger value reported by Picken may have been due to small
counterions (probably Na+) associated with the protein. This means that an increase
of 10-20-fold in protein concentration would probably still leave a positive filtration
pressure, for Picken reported hydrostatic pressures of 15 mm. Hg in the haemocoel,
and the value in a pathway as direct as that between the antennary artery* and
coelomosac may be even higher (as in the vertebrate glomerulus).
Concentration of protein by ultrafiltration should also generate a Donnan situation
in the coelomosac with the result that the concentration of diffusible cations (primarily
Na+) in the tubule should be lower than in the blood. Riegel has recently shown
(unpublished experiments) that the sodium concentration in the tubular fluid averaged
about 20% less than in blood taken from the ventral haemocoel. The discrepancy
might have been even more pronounced had it been possible to sample blood from the
peritubular vascular spaces in the coelomosac. Riegel's observation, which appears
to be incompatible with free filtration, is instead a predictable consequence of a large
filtration-fraction.
Thus the ability of the coelomosac to concentrate both endogenous and foreign
proteins points to this region as the site of formation of the ultrafiltrate. The data on
polymer excretion shows that a MW of above 150,000 is incompatible with filtration,
although molecular parameters other than size are undoubtedly involved. This is
* It has been reported (e.g. Peters, 1935 ; KUmmel, 1964) that the coelomosac is supplied by a branch
of the sternal artery. However, dye-perfusion experiments in this laboratory show that only the
posteromedial region of the green gland (comprising a portion of the labyrinth) is so supplied. Only
when a dye is perfused via the antennary artery does it appear in the coelomosac.
394
LEONARD B. KIRSCHNER AND STANLEY WAGNER
consistent with the blood-protein picture in decapods since the main constituent,
haemocyanin, is much too large to be filtered. However, the data presented here
indicate that further study will be required before we understand the mechanics of
filtrate formation in the antennal gland.
SUMMARY
1. Inulin (MW 5000) and two dextrans (MW 15,000-20,000 and 60,000-90,000)
appear in the urine of crayfish after injection into the blood. All three compounds are
more concentrated in the urine than in blood, indicating that water is reabsorbed from
a filtrate formed within the antennal gland.
2. Inulin and the low molecular weight dextran seem to be handled in the same
fashion since their urine/blood concentration ratios are about the same. However, the
high-molecular-weight dextran is excreted only about 70% as effectively as the other
pair. This suggests that filtration of polymers in the MW range 50,000-100,000 is
restrained.
3. Excretion of human serum albumin occurs but it is even more severely restricted
than the large dextran. Mammalian globulin (MW about 180,000) and crayfish blood
protein are not excreted in the urine.
4. Blood protein becomes concentrated in the coelomosac. Localization of fluorescein-labelled mammalian globulin shows that the peritubular cells in the coelomosac
are the main sites of protein accumulation. Concentration of blood protein in the
coelomosac suggests that filtration occurs in this region, but the intracellular location
suggests that the filtration mechanism differs from that in the vertebrate nephron.
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BAYLISS, L. E., KERRIDGE, P. M. T. & RUSSELL, D. S. (1933). Excretion of protein by the mammalian
kidney. J. Phytiol. 77, 386-98.
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BRAY, G. A. (i960). A simple efficient scintillator for counting aqueous solutions in a liquid scintillation
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PEARSE, A. G. E. (1961). Histochemistry. Boston: Little Brown and Co.
PETERS, H. (1935). tJber den Einfluss des Saltsgehaltes im Aussenmedium auf den Bau und die
Funktion der Exkretionsorgane dckapoder Crustacean. Z. Morph. Okol. Tiere, 30, 355—81.
RIEGEL, J. A. (1963). Micropuncture studies of chloride concentration and osmotic pressure in the
crayfish antennal gland. J. Exp. Biol. 40, 487—92.
RIBGEL, J. A. & KIRSCHNER, L. B. (i960). The excretion of inulin and glucose by the crayfish antennal
gland. Biol. Bull. 118, 296-307.
SMITH, H. W. (1951). The Kidney. Oxford University Press.
SOMOOYI, M. (1930). A method for the preparation of blood filtrates for the determination of blood
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Plate 1
Journal of Experimental Biology, Vol. 43, No. 2
tu
LEONARD B. KIRSCHNER AND STANLEY WAGNER
LFadng p. 395)
Filtration locus in crayfish antennal gland
395
EXPLANATION OF PLATE
Fig. 1. Evans's blue accumulation in the coelomosac. Experimental protocol is described in the text.
Fig. 2. Section through the labyrinth and tubule of an antennal gland removed 4 hr. after injection of
fluorescein-labelled globulin. The labyrinth comprises the outer border (upper edge) of the section;
the tubular tissue is more loosely arranged around larger lumina. The section is illuminated with
ultraviolet light. Magnification, x 100. The symbols delimit regions of the organ. Ib, labyrinth;
tu, tubule.
Fig. 3. Section through the coelomosac of the same organ shown in fig. 2. The coelomosac—tubule
border can be seen in the upper edge of the photomicrograph. Illumination and magnification as in
fig. 2. tu, Tubule; co, coelomosac; ptc, peritubular cells of coelomosac; lu, lumen of coelomosac.