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Ionic and osmotic regulation in the king crab and two other North Pacific crustaceans
Comp. Biochem. Physiol., 1970, Vol. 34, pp. 273 to 280. PergamonPress. Printed in Great Britain
IONIC AND OSMOTIC REGULATION IN THE KING CRAB AND TWO OTHER NORTH PACIFIC CRUSTACEANS W. C. MACKAY and C. L. PROSSER Department of Biology, Case Western Reserve University, Cleveland, Ohio; Department of Physiology and Biophysics, University of Illinois, Urbana, Illinois (Received 13 October 1969)
Abstract--1. King crab hemolymph was 5 to 20 m-osmoles hyperosmotic to the sea-water medium. The hemolymph of coon-striped shrimp and tanner crab was isosmotic to sea water. 2. Magnesium (36"9 m-moles/1.) in king crab hemolymph was lower and chloride (577 m-moles) higher than in sea water. 3. The coon-striped shrimp was the strongest ion regulator of the three species studied, the hemolymph concentrations of magnesium (5"8 m-moles) and chloride (466 m-moles) were lower, and calcium (12"3 m-moles) higher than in sea water. 4. Chloride (462 m-moles) was the only ion regulated by the tanner crab. INTRODUCTION THE hemolymph of most marine decapod crustaceans is isotonic with their environmental sea water but the ionic composition of their hemolymph can differ considerably from that of sea water (Krogh, 1939; Robertson, 1960; Prosser & Brown, 1961 ; Potts & Parry, 1963). In many marine crustaceans magnesium and sulphate are less concentrated in the hemolymph than in sea water. Hemolymph calcium concentration is normally greater than that of sea water while sodium, potassium and chloride concentrations are approximately equal to those found in sea water. Tissue analyses of decapods are less common than studies of their body fluids (Robertson, 1961 ; Dunham & Gainer, 1968). The most significant feature of the intracellular compartment is the high concentration of amino acids which make up a large proportion of the total intracellular osmolarity (Schoffeniels, 1967). The purpose of this study was to determine the extent and possible mechanisms of osmotic and ionic regulation in the king crab, Paralithodes camtschatica (Tilesius). Comparative data were also obtained for two other sublittoral decapods: the tanner crab, Chionoecetes tanneri (Rathburn) and the coon-striped shrimp, Pandalus hipsinotus.
Most previous studies on ionic balance in crustaceans have been on families in the sub-order Reptantia, section Brachyura. King crabs superficially resemble brachyuran spider crabs but actually belong to a different section. Paralithodes is in the family Lithodidae of the super-family Paguridea and section Anomura. The 273
274
W . C . MACKAY AND C. L. PROSSER
tanner crab is a typical brachyuran. T h e coon-striped shrimp is classified in the sub-order Natantia section Caridea. MATERIALS AND METHODS King crabs were obtained in the first week of July near the town of Kodiak, Alaska, by commercial crab traps. They were held on the ship in large tanks of circulating surface sea water or in traps over the side of the ship. The crabs were not fed for the 3-week duration of sampling. Some crabs were acclimated to diluted or concentrated sea water in aerated tanks and a few were placed near the mouth of a fresh-water stream for several days. Coonstriped shrimps and tanner crabs were caught by a small (6-ft dia.) otter trawl and held in tanks of circulating sea water for approximately 1 week before sampling. Hemolymph was obtained from the crabs by puncturing an arthroidal membrane on one of the legs and from the shrimp by puncture of the thorax in the region of the heart. Samples of muscle, green gland and gill weighing from 200 to 500 mg were taken from king crabs, blotted on paper towelling and weighed in tared vials. The tissues were then dried to constant weight at 60°C and the dry weights obtained. Dried tissue samples were crushed in vials with glass stirring rods and the ions were then extracted with regular mixing in 2 ml of distilled water for at least 24 hr. Samples of the extraction solution were then analyzed and ion concentrations in the tissue calculated. Osmolarities were determined on samples of 0"2 ml using an Advanced Standard Laboratory osmometer. Sodium and potassium were determined on a Baird Atomic model KY2 flame photometer and calcium on a Beckman DU Spectrophotometer using a hydrogen flame attachment. Magnesium was analysed by absorption spectrophotometry using a Perkin-Elmer atomic absorption unit. Chloride was determined by mercuric nitrate titration using diphenyl carbazine to indicate the end-point (Sigma Chloride Kit). RESULTS
Osmotic regulation T h e h e m o l y m p h of king crabs was found to be hyperosmotic to the environment over the environmental range of 800-983 m-osmoles (Fig. 1). T h e extent of the hypertonicity varied from 5 to 20 m-osmoles, the mean hypertonicity of the h e m o l y m p h to sea water being approximately 15 m-osmoles. W h e n some crabs were exposed to diluted sea water of 560 m-osmoles for a few hours, their abdomens became swollen and some died. T h e average h e m o l y m p h osmolarity of four crabs which did not die was 598 m-osmoles. T h e h e m o l y m p h of the other two sublittoral crustaceans examined, the coon-striped shrimp and the tanner crab, was isosmotic with the environment (Table 1).
Ionic composition of hemolymph T h e hypertonicity of king crab h e m o l y m p h can be accounted for by the higher concentration of sodium and chloride ions in the h e m o l y m p h than in sea water (Table 1). I n every case in which h e m o l y m p h and sea water were sampled simultaneously, the sodium concentration was 25-50 m-moles higher in the h e m o l y m p b than in sea water. T h e chloride concentration in the h e m o l y m p h for the same sample was from 50 to 100 m-moles higher than that of sea water. T h e concentration of sodium in the h e m o l y m p h of both the coon-striped shrimp and the tanner crab was not significantly different from that in sea water
CRUSTACEAN
275
ION REGULATION
while the chloride concentration in the hemolymph of both these animals was less than in sea water (Table 1). Tanner crabs, and to a lesser extent, the king crab, maintained higher potassium concentrations in the hemolymph than in sea water (Table 1). However, the tanner crab did not significantly regulate calcium and only slightly regulated magnesium (Table 1). The coon-striped shrimp concentrated calcium in the hemolymph while maintaining a very low concentration of magnesium in this fluid. The king crab regulated hemolymph magnesium below that of sea water (Table 1). / // //,
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Fro. 1. Relationship between osmotic concentration (milliosmolar) of king crab hemolymph and the sea water from which the animals were sampled. Dotted line represents an isosmotic relationship. Each symbol represents a sample from a single crab.
Ionic composition of king crab tissue The ratio of sodium to potassium in gill and green gland differed markedly from one another and from that in muscle (Table 2). Muscle contains very low concentrations of sodium but is rich in potassium, the Na to K ratio being 0"59. The high Na to K ratio (3"3) in gill could be due to sea water adhering to the gill lamellae or to the active uptake of sodium by the gills. Sodium and potassium concentrations were higher in the green gland than in muscle, but the ratio of Na to K (1.14) was only slightly higher than that found in muscle. The muscle ion concentration in m-moles/1, tissue water was calculated from the tissue ion concentration (m'moles/kg wet wt.) (Table 2). The water content of king crab muscle was 82.1 _+0.3 (S.E.) per cent (n = 20).
Hemolymph Sea water
Paralithodes camtschatica 933 + 5 (35) 902
896 + 16 (6) 907 461 + 9 (17) 452 + 3 (4)
395 + 10 (6) 380
409 + 7 (5) 393 + 19 (5) 417
403 + 11 (5) 420 _+23 (4)
Na
13"5 + 0"4 (16) 12'1 + 0"2 (4)
7"4 + 0'7 (6) 7"8
10"5 + 0"2 (5) 9"9 + 0"7 (5) 9"0
11.1 _+0"5 (5) 11 "4 + 1 '4 (4)
K
13 (2)
12"3 +_0"5 (6) 9"3
9'3 _+0"1 (5) 8"7 + 0"6 (5) 8"9
8"1 _+0-5 (5) 10"4 _+ 1 '4 (4)
Ca
Ion concentration (m-moles/L)
36"9 + 1-2 (13) 52'0
5"8 + 1"0 (6) 52.0
44"8 + 1"2 (5) 45"2 _+3'5 (5) 52"5
Mg
577 + 12 (13) 494 + 4 (5)
466 + 9 (6) 493
462 + 2 (5) 456 _+6 (5) 490
495 + 3 (5) 482 + 23 (4)
CI
AND SEA W A T E R F R O M W H I C H T H E A N I M A L S W E R E S A M P L E D
T h e first n u m b e r in each column is the mean of the samples followed by the standard error of the m e a n (S.E.) and in brackets the n u m b e r of individuals in the sample.
Hemolymph Sea water
862 + 2 (5)
Hemolymph Urine Sea water 860
998 + 9 (4) 997 + 2 (5)
Osmolarity (m-osmoles)
Hemolymph Urine
Fluid
AND O S M O T I C C O N C E N T R A T I O N OF H E M O L Y M P H
Pandalus hipsinotus
Chionoecetes tanneri
Species
TABLE 1--IoNIC
t~
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(3
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277
CRUSTACEAN I O N REGULATION TABLE 2--ION
CONCENTRATION I N K I N G CRAB MUSCLEj GREEN GLAND AND GILLS I N
m-moles/kg
W E T W T . OF THE TISSUE
Ion concentration Tissue Whole muscle (m-moles/kg wet wt.) Whole muscle (m-moles/1. tissue water) Gill (m-moles/kg wet wt.) Grand gland (m-moles/kg wet wt.)
Na
K
Ca
Mg
62 (6) 76 195 (6) 144 (4)
105 (8) 128 59 (6) 126 (4)
9 (1) 11 11 (3)
8 (2) 10 8 (3)
The upper number in each case is the mean. The number of individuals in the sample is included in brackets below. Muscle ion concentrations in m-moles/1, tissue water were calculated from whole muscle ion concentrations and water content. The green gland of the king crab is located at the base of the second antenna and opens to the exterior through an inconspicuous pore on the ventral side of the base of the antenna. The gland itself is small in relation to the size of the crab, and the bladder is represented by a slight enlargement of the small duct leading to the exterior. Although several attempts were made, no urine could be collected from the king crab because of the location of the external opening, the small duct and poorly developed bladder. Urine was easily obtained from the tanner crab. The ionic composition of tanner crab urine was similar to that of the hemolymph (Table 1). DISCUSSION Since no successful urine collections could be made, all of the data concerning osmotic and ionic regulation in the king crab were obtained by sampling hemolymph, sea water and by tissue analysis. Many marine crustaceans living in full strength sea water have been found to be isosmotic with the sea water (Robertson, 1960). However, the king crab was hyperosmotic to sea water at all salinities tested. The inability of this animal to maintain volume regulation or to live even for a short time at 560 m-osmoles attests to its stenohalinity. The mechanism by which hypertonicity is maintained has not been elucidated. It is known that an increase in the osmotic concentration of the hemolymph is part of the mechanism for water uptake during molting (Passano, 1960). The osmotic pressure of the hemolymph increases rapidly a short time before molting and remains above the osmotic pressure of sea water until the skeleton is hardened (Baumberger & Olmsted, 1928). King crabs molt in early summer. Most of the animals used for this study had molted several weeks prior to sampling. However, the hemolymph of crabs which had not molted for 1 year was also hyperosmotic to the sea water in which they were held.
0"98 0"98 1"11 1.02 1 "01 0"83 1'07 1-11 1-13 1.04 1"09 1-04 0-85 1"19
Chionoecetes Cancer Carcinus
Pachygrapsus Uca Homarus
Pandalus Palaemon Palinurus 0"95 0"85 1"05
1 '17 1"02 1"21 0"95 1 '23 1"22 1"00 0'86 0'78 1"27 1"30
1' 12
K
1 '32 1'05 1.33
1 "04 1"13 1"27 1'23 1 "14 1"33 1"68 1"51 1"37 1.22 1'38
--
Ca
0' 11 0-20 0'32
0'85 0"41 0"36 0-42 0-56 0"52 0"13 0'14 0'17 1"00 0"86
0-71
Mg
0'95 0"85 1.03
0'94 0'88 1"00 0'98 -0"93 0"94 1'00 0'97 0-93 1.06
1" 17
C1
-0'10 0"77
-0-67 0"57 --1"91 -0"31 0"67 1"00 0"54
--
SO,
Reference
Present study Parry, 1954 R o b e r t s o n , 1949
Present study Nicol, 1967 W e b b , 1940 Shaw, 1955 Prosser et al., 1955 G r e e n et aL, 1959 Burger, 1957 R o b e r t s o n , 1949 R o b e r t s o n , 1949 R o b e r t s o n , 1949 R o b e r t s o n , 1949
Present study
OF I O N R E G U L A T I O N OF S O M E M A R I N E CRU ST A CE A N S I N F U L L SEA W A T E R
T h e c o n c e n t r a t i o n of t h e ion in the h e m o l y m p h has b e e n divided b y the c o n c e n t r a t i o n of t h a t ion f o u n d in the sea water f r o m w h i c h the a n i m a l was sampled.
Nephros Lithodes .~Vlaja
1 "02
Na
Paralithodes
Species
TABLE 3--COMPARISON
b~
ct~ t~
P
ta
P
OO
CRUSTACEAN I O N R E G U L A T I O N
279
T h e tanner crab and the coon-striped shrimp are typical of other stenohaline marine crustaceans in that their hemolymph and in the case of the tanner crab, its urine, are isosmotic with sea water. T h e hypertonicity of king crab hemolymph can be accounted for by sodium and chloride ions. Since the concentration of both of these ions in the hemolymph is against a concentration gradient, some tissue, probably the gills, must be actively involved in taking up sodium, chloride or both. A few measurements of potentials across the gills (3 to 5 mV inside positive) support the active uptake of sodium. T h e ions in the hemolymph of king crab are regulated to approximately the same extent as in other marine crustaceans. Potassium is somewhat more concentrated and magnesium is considerably less concentrated in the hemolymph than in sea water. T h e high concentration of both sodium and potassium in the green gland may indicate excretion of these ions by this tissue. Since urine samples could not be collected, the role of the green gland in ion regulation could not be ascertained. Because the hemolymph is hypertonic to sea water one would expect the green gland to function in the excretion of water gained osmotically from sea water. T he green gland may function to excrete other compounds such as acid. Howell (unpublished data) has observed that king crabs are able to lower the acidity of several gallons of sea water by more than one pH unit in a few hours without affecting the hemolymph pH. T he site of secretion and the nature of the acid were not determined. Comparisons of hemolymph ion concentrations among crustaceans may be made by dividing the concentration of an ion in the hemolymph by its concentration in sea water. This has been done for the data obtained in this study and for data on other marine crustaceans in sea water (Table 3). When such ratios are compared it becomes apparent that the relative concentration of chloride in the hemolymph of Paralithodes is much greater than that found in other marine crustaceans. T he ratios obtained for the tanner crab, Chionoecetes, are similar to those of other brachyuran crabs (Table 3). However, the coon-striped shrimp, Pandulus, shows a greater regulation of magnesium than any of the other crustaceans shown (Table 3). Acknowledgements--This work was carried out aboard the R.V. Alpha Helix, of the University. of California, San Diego, during Phase C of the Bering Sea Expedition and was supported by the National Science Foundation, U.S.A. One of the authors (W. C. M.) was also supported by N.I.H. Comparative Physiology Training Grant No. GM-01699-03. REFERENCES
BAUMBERGERJ. P. & OLMSTEDJ. M. D. (1928) Changes in the osmotic pressure and water content of crabs during the molt cycle. Physiol. Zo6l. 1,531-544. BURCER J. W. (1957) The general form of excretion in the lobster, Homarus. Biol. Bull. 113, 207-223. DUNHAM P. B. & GAINERH. (1968) The distribution of inorganic ions in lobster muscle. Biochim. biophys. Acta 150,488-499. GREEN J. W., HARSCHM., BARRL. & PROSSERC. L. (1959) Ionic regulation in Uca. Biol. Bull. 116, 76-87.
280
W. C. MACKAYAND C. L. PROSSER
KROGH A. (1939) Osmotic Regulation in Aquatic Animals. Dover, New York. NmoL J. A. C. (1967) The Biology of Marine Animals, 2nd edn. Pitman, London. PARRY G. (1954) Ionic regulation in the prawn, Palaemon serratus, ft. exp. Biol. 31, 601-613. PASSANOL. M. (1960) Molting and its control. In Physiology of Crustacea, Vol. 1, Metabolism and Growth (Edited by WATERMANT. H.), Chapt. 5. Academic Press, New York. POTTS W. T. W. & PARRY G. (1963) Osmotic and Ionic Regulation in Animals. Pergamon, New York. PROSSER C. L. & BROWN F. A. (1961) Comparative Animal Physiology, 2nd edn. Saunders, Philadelphia. PROSSER C. L., GI~_~_~J. W. & CHOW T. J. (1955) Ionic balance in Pachygrapsus. Biol. Bull. 109, 99-107. ROBERTSON J. D. (1961) Studies on the chemical composition of muscle t i s s u e - - I I . T h e abdominal flexor muscles of the lobster, Nephrops norvegicus, ft. exp. Biol. 38, 707-728. ROBERTSON,J. D. (1960) Osmotic and ionic regulation. In Physiology of Crustacea, Vol. I, Metabolism and Growth. (Edited by WATERMANT. H.), Chapt. 9. Academic Press, New York. ROBERTSON J. D. (1949) Ionic Regulation in Marine Invertebrates. ft. exp. Biol. 26, 182-200. SCHOFF~NmLS E. (1967) Cellular Aspects of Membrane Permeability. Pergamon, Oxford. SHAW J. (1955) Ionic regulation in muscle fibres, Carcinus. ft. exp. Biol. 32, 383-396. WEBB D. A. (1940) Osmotic and ionic regulation in Carcinus. Proc. R. Soc. (B) 129, 107136.
Key Word Index---Ionic regulation; osmotic regulation; crustacean tissue ion concentration; intracellular ions.
IONIC AND OSMOTIC REGULATION IN THE KING CRAB AND TWO OTHER NORTH PACIFIC CRUSTACEANS W. C. MACKAY and C. L. PROSSER Department of Biology, Case Western Reserve University, Cleveland, Ohio; Department of Physiology and Biophysics, University of Illinois, Urbana, Illinois (Received 13 October 1969)
Abstract--1. King crab hemolymph was 5 to 20 m-osmoles hyperosmotic to the sea-water medium. The hemolymph of coon-striped shrimp and tanner crab was isosmotic to sea water. 2. Magnesium (36"9 m-moles/1.) in king crab hemolymph was lower and chloride (577 m-moles) higher than in sea water. 3. The coon-striped shrimp was the strongest ion regulator of the three species studied, the hemolymph concentrations of magnesium (5"8 m-moles) and chloride (466 m-moles) were lower, and calcium (12"3 m-moles) higher than in sea water. 4. Chloride (462 m-moles) was the only ion regulated by the tanner crab. INTRODUCTION THE hemolymph of most marine decapod crustaceans is isotonic with their environmental sea water but the ionic composition of their hemolymph can differ considerably from that of sea water (Krogh, 1939; Robertson, 1960; Prosser & Brown, 1961 ; Potts & Parry, 1963). In many marine crustaceans magnesium and sulphate are less concentrated in the hemolymph than in sea water. Hemolymph calcium concentration is normally greater than that of sea water while sodium, potassium and chloride concentrations are approximately equal to those found in sea water. Tissue analyses of decapods are less common than studies of their body fluids (Robertson, 1961 ; Dunham & Gainer, 1968). The most significant feature of the intracellular compartment is the high concentration of amino acids which make up a large proportion of the total intracellular osmolarity (Schoffeniels, 1967). The purpose of this study was to determine the extent and possible mechanisms of osmotic and ionic regulation in the king crab, Paralithodes camtschatica (Tilesius). Comparative data were also obtained for two other sublittoral decapods: the tanner crab, Chionoecetes tanneri (Rathburn) and the coon-striped shrimp, Pandalus hipsinotus.
Most previous studies on ionic balance in crustaceans have been on families in the sub-order Reptantia, section Brachyura. King crabs superficially resemble brachyuran spider crabs but actually belong to a different section. Paralithodes is in the family Lithodidae of the super-family Paguridea and section Anomura. The 273
274
W . C . MACKAY AND C. L. PROSSER
tanner crab is a typical brachyuran. T h e coon-striped shrimp is classified in the sub-order Natantia section Caridea. MATERIALS AND METHODS King crabs were obtained in the first week of July near the town of Kodiak, Alaska, by commercial crab traps. They were held on the ship in large tanks of circulating surface sea water or in traps over the side of the ship. The crabs were not fed for the 3-week duration of sampling. Some crabs were acclimated to diluted or concentrated sea water in aerated tanks and a few were placed near the mouth of a fresh-water stream for several days. Coonstriped shrimps and tanner crabs were caught by a small (6-ft dia.) otter trawl and held in tanks of circulating sea water for approximately 1 week before sampling. Hemolymph was obtained from the crabs by puncturing an arthroidal membrane on one of the legs and from the shrimp by puncture of the thorax in the region of the heart. Samples of muscle, green gland and gill weighing from 200 to 500 mg were taken from king crabs, blotted on paper towelling and weighed in tared vials. The tissues were then dried to constant weight at 60°C and the dry weights obtained. Dried tissue samples were crushed in vials with glass stirring rods and the ions were then extracted with regular mixing in 2 ml of distilled water for at least 24 hr. Samples of the extraction solution were then analyzed and ion concentrations in the tissue calculated. Osmolarities were determined on samples of 0"2 ml using an Advanced Standard Laboratory osmometer. Sodium and potassium were determined on a Baird Atomic model KY2 flame photometer and calcium on a Beckman DU Spectrophotometer using a hydrogen flame attachment. Magnesium was analysed by absorption spectrophotometry using a Perkin-Elmer atomic absorption unit. Chloride was determined by mercuric nitrate titration using diphenyl carbazine to indicate the end-point (Sigma Chloride Kit). RESULTS
Osmotic regulation T h e h e m o l y m p h of king crabs was found to be hyperosmotic to the environment over the environmental range of 800-983 m-osmoles (Fig. 1). T h e extent of the hypertonicity varied from 5 to 20 m-osmoles, the mean hypertonicity of the h e m o l y m p h to sea water being approximately 15 m-osmoles. W h e n some crabs were exposed to diluted sea water of 560 m-osmoles for a few hours, their abdomens became swollen and some died. T h e average h e m o l y m p h osmolarity of four crabs which did not die was 598 m-osmoles. T h e h e m o l y m p h of the other two sublittoral crustaceans examined, the coon-striped shrimp and the tanner crab, was isosmotic with the environment (Table 1).
Ionic composition of hemolymph T h e hypertonicity of king crab h e m o l y m p h can be accounted for by the higher concentration of sodium and chloride ions in the h e m o l y m p h than in sea water (Table 1). I n every case in which h e m o l y m p h and sea water were sampled simultaneously, the sodium concentration was 25-50 m-moles higher in the h e m o l y m p b than in sea water. T h e chloride concentration in the h e m o l y m p h for the same sample was from 50 to 100 m-moles higher than that of sea water. T h e concentration of sodium in the h e m o l y m p h of both the coon-striped shrimp and the tanner crab was not significantly different from that in sea water
CRUSTACEAN
275
ION REGULATION
while the chloride concentration in the hemolymph of both these animals was less than in sea water (Table 1). Tanner crabs, and to a lesser extent, the king crab, maintained higher potassium concentrations in the hemolymph than in sea water (Table 1). However, the tanner crab did not significantly regulate calcium and only slightly regulated magnesium (Table 1). The coon-striped shrimp concentrated calcium in the hemolymph while maintaining a very low concentration of magnesium in this fluid. The king crab regulated hemolymph magnesium below that of sea water (Table 1). / // //,
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Fro. 1. Relationship between osmotic concentration (milliosmolar) of king crab hemolymph and the sea water from which the animals were sampled. Dotted line represents an isosmotic relationship. Each symbol represents a sample from a single crab.
Ionic composition of king crab tissue The ratio of sodium to potassium in gill and green gland differed markedly from one another and from that in muscle (Table 2). Muscle contains very low concentrations of sodium but is rich in potassium, the Na to K ratio being 0"59. The high Na to K ratio (3"3) in gill could be due to sea water adhering to the gill lamellae or to the active uptake of sodium by the gills. Sodium and potassium concentrations were higher in the green gland than in muscle, but the ratio of Na to K (1.14) was only slightly higher than that found in muscle. The muscle ion concentration in m-moles/1, tissue water was calculated from the tissue ion concentration (m'moles/kg wet wt.) (Table 2). The water content of king crab muscle was 82.1 _+0.3 (S.E.) per cent (n = 20).
Hemolymph Sea water
Paralithodes camtschatica 933 + 5 (35) 902
896 + 16 (6) 907 461 + 9 (17) 452 + 3 (4)
395 + 10 (6) 380
409 + 7 (5) 393 + 19 (5) 417
403 + 11 (5) 420 _+23 (4)
Na
13"5 + 0"4 (16) 12'1 + 0"2 (4)
7"4 + 0'7 (6) 7"8
10"5 + 0"2 (5) 9"9 + 0"7 (5) 9"0
11.1 _+0"5 (5) 11 "4 + 1 '4 (4)
K
13 (2)
12"3 +_0"5 (6) 9"3
9'3 _+0"1 (5) 8"7 + 0"6 (5) 8"9
8"1 _+0-5 (5) 10"4 _+ 1 '4 (4)
Ca
Ion concentration (m-moles/L)
36"9 + 1-2 (13) 52'0
5"8 + 1"0 (6) 52.0
44"8 + 1"2 (5) 45"2 _+3'5 (5) 52"5
Mg
577 + 12 (13) 494 + 4 (5)
466 + 9 (6) 493
462 + 2 (5) 456 _+6 (5) 490
495 + 3 (5) 482 + 23 (4)
CI
AND SEA W A T E R F R O M W H I C H T H E A N I M A L S W E R E S A M P L E D
T h e first n u m b e r in each column is the mean of the samples followed by the standard error of the m e a n (S.E.) and in brackets the n u m b e r of individuals in the sample.
Hemolymph Sea water
862 + 2 (5)
Hemolymph Urine Sea water 860
998 + 9 (4) 997 + 2 (5)
Osmolarity (m-osmoles)
Hemolymph Urine
Fluid
AND O S M O T I C C O N C E N T R A T I O N OF H E M O L Y M P H
Pandalus hipsinotus
Chionoecetes tanneri
Species
TABLE 1--IoNIC
t~
(3
(3
tO ,.q
277
CRUSTACEAN I O N REGULATION TABLE 2--ION
CONCENTRATION I N K I N G CRAB MUSCLEj GREEN GLAND AND GILLS I N
m-moles/kg
W E T W T . OF THE TISSUE
Ion concentration Tissue Whole muscle (m-moles/kg wet wt.) Whole muscle (m-moles/1. tissue water) Gill (m-moles/kg wet wt.) Grand gland (m-moles/kg wet wt.)
Na
K
Ca
Mg
62 (6) 76 195 (6) 144 (4)
105 (8) 128 59 (6) 126 (4)
9 (1) 11 11 (3)
8 (2) 10 8 (3)
The upper number in each case is the mean. The number of individuals in the sample is included in brackets below. Muscle ion concentrations in m-moles/1, tissue water were calculated from whole muscle ion concentrations and water content. The green gland of the king crab is located at the base of the second antenna and opens to the exterior through an inconspicuous pore on the ventral side of the base of the antenna. The gland itself is small in relation to the size of the crab, and the bladder is represented by a slight enlargement of the small duct leading to the exterior. Although several attempts were made, no urine could be collected from the king crab because of the location of the external opening, the small duct and poorly developed bladder. Urine was easily obtained from the tanner crab. The ionic composition of tanner crab urine was similar to that of the hemolymph (Table 1). DISCUSSION Since no successful urine collections could be made, all of the data concerning osmotic and ionic regulation in the king crab were obtained by sampling hemolymph, sea water and by tissue analysis. Many marine crustaceans living in full strength sea water have been found to be isosmotic with the sea water (Robertson, 1960). However, the king crab was hyperosmotic to sea water at all salinities tested. The inability of this animal to maintain volume regulation or to live even for a short time at 560 m-osmoles attests to its stenohalinity. The mechanism by which hypertonicity is maintained has not been elucidated. It is known that an increase in the osmotic concentration of the hemolymph is part of the mechanism for water uptake during molting (Passano, 1960). The osmotic pressure of the hemolymph increases rapidly a short time before molting and remains above the osmotic pressure of sea water until the skeleton is hardened (Baumberger & Olmsted, 1928). King crabs molt in early summer. Most of the animals used for this study had molted several weeks prior to sampling. However, the hemolymph of crabs which had not molted for 1 year was also hyperosmotic to the sea water in which they were held.
0"98 0"98 1"11 1.02 1 "01 0"83 1'07 1-11 1-13 1.04 1"09 1-04 0-85 1"19
Chionoecetes Cancer Carcinus
Pachygrapsus Uca Homarus
Pandalus Palaemon Palinurus 0"95 0"85 1"05
1 '17 1"02 1"21 0"95 1 '23 1"22 1"00 0'86 0'78 1"27 1"30
1' 12
K
1 '32 1'05 1.33
1 "04 1"13 1"27 1'23 1 "14 1"33 1"68 1"51 1"37 1.22 1'38
--
Ca
0' 11 0-20 0'32
0'85 0"41 0"36 0-42 0-56 0"52 0"13 0'14 0'17 1"00 0"86
0-71
Mg
0'95 0"85 1.03
0'94 0'88 1"00 0'98 -0"93 0"94 1'00 0'97 0-93 1.06
1" 17
C1
-0'10 0"77
-0-67 0"57 --1"91 -0"31 0"67 1"00 0"54
--
SO,
Reference
Present study Parry, 1954 R o b e r t s o n , 1949
Present study Nicol, 1967 W e b b , 1940 Shaw, 1955 Prosser et al., 1955 G r e e n et aL, 1959 Burger, 1957 R o b e r t s o n , 1949 R o b e r t s o n , 1949 R o b e r t s o n , 1949 R o b e r t s o n , 1949
Present study
OF I O N R E G U L A T I O N OF S O M E M A R I N E CRU ST A CE A N S I N F U L L SEA W A T E R
T h e c o n c e n t r a t i o n of t h e ion in the h e m o l y m p h has b e e n divided b y the c o n c e n t r a t i o n of t h a t ion f o u n d in the sea water f r o m w h i c h the a n i m a l was sampled.
Nephros Lithodes .~Vlaja
1 "02
Na
Paralithodes
Species
TABLE 3--COMPARISON
b~
ct~ t~
P
ta
P
OO
CRUSTACEAN I O N R E G U L A T I O N
279
T h e tanner crab and the coon-striped shrimp are typical of other stenohaline marine crustaceans in that their hemolymph and in the case of the tanner crab, its urine, are isosmotic with sea water. T h e hypertonicity of king crab hemolymph can be accounted for by sodium and chloride ions. Since the concentration of both of these ions in the hemolymph is against a concentration gradient, some tissue, probably the gills, must be actively involved in taking up sodium, chloride or both. A few measurements of potentials across the gills (3 to 5 mV inside positive) support the active uptake of sodium. T h e ions in the hemolymph of king crab are regulated to approximately the same extent as in other marine crustaceans. Potassium is somewhat more concentrated and magnesium is considerably less concentrated in the hemolymph than in sea water. T h e high concentration of both sodium and potassium in the green gland may indicate excretion of these ions by this tissue. Since urine samples could not be collected, the role of the green gland in ion regulation could not be ascertained. Because the hemolymph is hypertonic to sea water one would expect the green gland to function in the excretion of water gained osmotically from sea water. T he green gland may function to excrete other compounds such as acid. Howell (unpublished data) has observed that king crabs are able to lower the acidity of several gallons of sea water by more than one pH unit in a few hours without affecting the hemolymph pH. T he site of secretion and the nature of the acid were not determined. Comparisons of hemolymph ion concentrations among crustaceans may be made by dividing the concentration of an ion in the hemolymph by its concentration in sea water. This has been done for the data obtained in this study and for data on other marine crustaceans in sea water (Table 3). When such ratios are compared it becomes apparent that the relative concentration of chloride in the hemolymph of Paralithodes is much greater than that found in other marine crustaceans. T he ratios obtained for the tanner crab, Chionoecetes, are similar to those of other brachyuran crabs (Table 3). However, the coon-striped shrimp, Pandulus, shows a greater regulation of magnesium than any of the other crustaceans shown (Table 3). Acknowledgements--This work was carried out aboard the R.V. Alpha Helix, of the University. of California, San Diego, during Phase C of the Bering Sea Expedition and was supported by the National Science Foundation, U.S.A. One of the authors (W. C. M.) was also supported by N.I.H. Comparative Physiology Training Grant No. GM-01699-03. REFERENCES
BAUMBERGERJ. P. & OLMSTEDJ. M. D. (1928) Changes in the osmotic pressure and water content of crabs during the molt cycle. Physiol. Zo6l. 1,531-544. BURCER J. W. (1957) The general form of excretion in the lobster, Homarus. Biol. Bull. 113, 207-223. DUNHAM P. B. & GAINERH. (1968) The distribution of inorganic ions in lobster muscle. Biochim. biophys. Acta 150,488-499. GREEN J. W., HARSCHM., BARRL. & PROSSERC. L. (1959) Ionic regulation in Uca. Biol. Bull. 116, 76-87.
280
W. C. MACKAYAND C. L. PROSSER
KROGH A. (1939) Osmotic Regulation in Aquatic Animals. Dover, New York. NmoL J. A. C. (1967) The Biology of Marine Animals, 2nd edn. Pitman, London. PARRY G. (1954) Ionic regulation in the prawn, Palaemon serratus, ft. exp. Biol. 31, 601-613. PASSANOL. M. (1960) Molting and its control. In Physiology of Crustacea, Vol. 1, Metabolism and Growth (Edited by WATERMANT. H.), Chapt. 5. Academic Press, New York. POTTS W. T. W. & PARRY G. (1963) Osmotic and Ionic Regulation in Animals. Pergamon, New York. PROSSER C. L. & BROWN F. A. (1961) Comparative Animal Physiology, 2nd edn. Saunders, Philadelphia. PROSSER C. L., GI~_~_~J. W. & CHOW T. J. (1955) Ionic balance in Pachygrapsus. Biol. Bull. 109, 99-107. ROBERTSON J. D. (1961) Studies on the chemical composition of muscle t i s s u e - - I I . T h e abdominal flexor muscles of the lobster, Nephrops norvegicus, ft. exp. Biol. 38, 707-728. ROBERTSON,J. D. (1960) Osmotic and ionic regulation. In Physiology of Crustacea, Vol. I, Metabolism and Growth. (Edited by WATERMANT. H.), Chapt. 9. Academic Press, New York. ROBERTSON J. D. (1949) Ionic Regulation in Marine Invertebrates. ft. exp. Biol. 26, 182-200. SCHOFF~NmLS E. (1967) Cellular Aspects of Membrane Permeability. Pergamon, Oxford. SHAW J. (1955) Ionic regulation in muscle fibres, Carcinus. ft. exp. Biol. 32, 383-396. WEBB D. A. (1940) Osmotic and ionic regulation in Carcinus. Proc. R. Soc. (B) 129, 107136.
Key Word Index---Ionic regulation; osmotic regulation; crustacean tissue ion concentration; intracellular ions.