Microscopy AND Microanalysis MICROSCOPY SOCIETY OF AMERICA 2005

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1 Microsc. Microanal. 11, 18 36, 2005 DOI: /S Microscopy AND Microanalysis MICROSCOPY SOCIETY OF AMERICA 2005 The Decapod Crustacean Circulatory System: A Case That Is neither Open nor Closed Iain J. McGaw 1,2 * 1 Department of Biological Sciences, University of Nevada, Las Vegas, NV 89154, USA 2 Bamfield Marine Sciences Centre, Bamfield, British Columbia V0R 1B0, Canada Abstract: Historically, the decapod crustacean circulatory system has been classed as open. However, recent work on the blue crab, Callinectes sapidus, suggests the circulatory system may be more complex than previously described. Corrosion casting techniques were refined and used to map the circulatory system of a variety of crab species ~order: Decapoda; family: Cancridae! to determine if the complexity observed in the blue crab was present in other species. Seven arteries arose from the single chambered heart. The anterior aorta, the paired anterolateral arteries, and the paired hepatic arteries exited from the anterior aspect of the heart. The small-diameter posterior aorta exited posteriorly from the heart. Exiting from the ventral surface of the heart, the sternal artery branched to supply the legs and mouthparts of the crab. These arteries were more complex than previously described, with arterioles perfusing all areas of the body. The arterioles split into fine capillary-like vessels. Most of these capillaries were blind ending. However, in several areas ~antennal gland, supraesophageal ganglion! complete capillary beds were present. After passing through the capillary-like vessels, blood drained into a series of sinuses. However, rather than being arbitrary spaces as previously described, scanning electron micrographs showed the sinuses to be distinct units. Most of the sinuses formed a series of flattened membrane-bound lacunae. This complexity may qualify the decapod crustacean circulatory system as one that is partially closed rather than open. Key words: artery, crab, circulation, cardiovascular, heart, sinus, corrosion casting, scanning electron microscopy INTRODUCTION The decapod crustacean circulatory system is traditionally classed as an open system. It consists of a single-chambered ventricle that is suspended in a primer chamber, the pericardial sinus ~Maynard, 1960!. Hemolymph ~blood! is pumped out into seven arteries ~five arterial systems!.three arterial systems deliver hemolymph anteriorly. The anterior aorta supplies the eyestalks, antennae, and supraesophageal ganglion of the crab. The gonads, antennal glands, and anterior regions of the digestive system are supplied by the paired anterolateral arteries, and the paired hepatic arteries branch within the hepatopancreas. Exiting posteriorly from the heart is the posterior aorta; this small artery and associated branches perfuse the abdomen and hindgut. The largest vessel, the sternal artery, exits the heart ventrally and divides to supply the limbs and mouthparts ~Pearson, 1908; McLaughlin, 1983!. Each artery is valved at its origin ~Alexandrowicz, 1932! and opening or closing of the valves controls hemolymph flow into each arterial system. All arteries branch into smaller arteries and fine capillary-like Received September 3, 2003; accepted February 5, * imcgaw@ccmail.nevada.edu vessels that ramify within the tissues, where gas, nutrient, and waste exchange take place. However, some of these small vessels may actually form true capillary beds ~Sandeman, 1967!. Nevertheless, decapod crustaceans lack a complete venous system. Once hemolymph has bathed the organs it drains into interstitial lacunae, ~small irregular spaces between tissues!, then into sinuses, which are large irregular intertissue spaces ~Maynard, 1960! that are often so poorly defined that they almost defy successful demonstration ~Pyle & Cronin, 1950!. The hemolymph from these sinuses eventually collects in the large ventral thoracic sinus below the heart, which extends into five branchial sinuses. These empty into the infrabranchial sinus, which extends along the base of the gills. From here, hemolymph flows through the gills by way of the afferent and efferent branchial veins and through the lamellae, where it is oxygenated. Hemolymph returns to the pericardial sinus via the branchiocardiac veins and then flows into the heart through three pairs of ostia ~Maynard, 1960; McLaughlin, 1983!. During the last decade, physiological aspects of the decapod crustacean cardiovascular system have received extensive investigation ~see McMahon & Burnett, 1990; McGaw et al., 1994a; McMahon et al., 1997; McMahon, 1999; Wilkens, 1999!. The system is now recognized as being relatively complex, with control mechanisms and

2 The Crustacean Circulatory System 19 operating pressures that rival some of the simple vertebrate closed systems ~McMahon & Burnett, 1990; McGaw & Mc- Mahon, 1999; Wilkens, 1999!. Recent work on the gross anatomy of the blue crab ~Callinectes sapidus! circulatory system ~McGaw & Reiber, 2002! substantiates these physiological processes. It shows that the circulatory system is much more complex than both the classical works ~Haeckel, 1857; Claus, 1884; Bouvier, 1891; Pearson, 1908; Baumann, 1921; Brody & Perkins, 1930! and some of the more recent, highly cited articles ~McLaughlin, 1983!, suggest. The complex circulatory system of C. sapidus may now be considered to be partially closed ~McGaw & Reiber, 2002!. However, it is unknown if the intricacy observed in C. sapidus is confined to this species. Therefore, the aim of the present study was to refine corrosion casting techniques and to use these to map the circulatory system, to provide detailed photographs showing the complexity of the system across several species of decapod crustaceans. MATERIALS AND METHODS Brachyuran crabs ~Family, Cancridae! were collected along the coast of California ~Cancer antennarius, Cancer anthonyi, Cancer gracilis, Cancer productus! or purchased from commercial suppliers ~Cancer magister, Cancer irroratus!. They were held in a recirculating artificial seawater system ~Instant Ocean! at a temperature of C and a salinity of 32 ppt and fed chopped fish twice weekly prior to use. Batsons Monomer resin No. 17 ~Polysciences Inc., Warrington, PA! was used to map the circulatory system. Crabs were anesthetized by submerging them in iced water for 30 min. A small hole was then drilled into the carapace directly above the heart and a polyethylene catheter ~PE50 to PE90! was implanted into the pericardial sinus of the crab. A syringe pump was used to infuse the Monomer that had been thinned with methacrylate acid methyl ester ~6:1! into the pericardial sinus. The crabs were kept immersed in a tank of seawater while infusion volumes of approximately 20% of the wet mass of the animal were injected at 2 ml/min. The tips of two of the legs were cut off to allow hemolymph to escape. Infusion pressures were maintained at or below 15 mmhg ~within physiological range! to prevent arterial damage caused by overfilling and excess pressure. Individual vessels were also cannulated and perfused with monomer at a rate of 1 ml/min, using the methods described above. Following injection of the Batsons Monomer, the crabs were stored at 58C for 12 h to allow the resin to cure. The soft tissue was macerated over 3 days in a saturated potassium hydroxide solution. The remaining carapace and chitinous tissues were then dried at room temperature and subsequently dissolved in concentrated hydrochloric acid ~1 M! for 12 h. Finally the cast was washed in distilled water to remove any remaining material. This procedure left a corrosion cast of the circulatory system with all tissues removed, thus avoiding the need for removal of material manually, which could damage the cast. The larger arterial systems were photographed with an SLR camera and 105-mm macro lens. Specimens were also photographed at higher magnifications using a stereoscopic dissecting microscope ~Leitz! at magnification. Fine detail of the arteries were examined using a scanning electron microscope. The casts were sputter-coated with gold and examined with a JEOL scanning electron microscope ~JSM-5600!. The final image quality was optimized using Adobe Photoshop. RESULTS Seven arteries ~five arterial systems! originate from the brachyuran crab heart. Arteries are defined as large vessels of circular diameter that supply specific areas of the body. The arteries branch into small diameter capillary-like vessels ~,50 mm! that perfuse an organ or organ system. From here, hemolymph collects in sinuses, which are irregular intertissue spaces ~Maynard, 1960!. Many sinuses drain into lacunae, which are smaller, flattened channels, with definite structure. Several veins return hemolymph back to the heart. By definition, the veins differ from sinuses because they are large vessels with a circular cross section that function solely as blood-flow channels ~Maynard, 1960!. Each of the vessel systems is discussed in turn. Anterior Aorta Complex The anterior aorta ~AA! exits from the medial dorsal anterior aspect of the heart ~Fig. 1a!. It continues anteriorly over the gonads as well as the pyloric and cardiac stomachs. It then dips ventrally toward the rostrum on the anterior edge of the carapace. Small side branches ~arrow! emanate along the entire length of the anterior aorta, supplying tissues close to the main artery. Most of these delicate vessels were lost on the present cast. At the end of the anterior aorta is an enlarged region, the cor frontale ~CF!. Three arteries extend from the cor frontale. The small cerebral artery ~CA! branches anteriorly, splitting into numerous smaller vessels ~on!. These extend around the optic and oculomotor neuropiles in the supraesophageal ganglion ~Fig. 1b!. The cerebral artery continues its course anteriorly and splits terminally ~ol!; these small vessels supply the olfactory lobe. The paired optic arteries ~OA!, which supply hemolymph to the eyes, exit at a 458 angle to the anterior aorta. One lateral optic artery ~LOA! protrudes from each optic artery close to the junction with the eye ~Fig. 1a!. The lateral optic arteries are situated 1808 relative to the optic arteries, and run back toward the midline of the carapace. They supply hemolymph to the retractor and extensor muscles of the eyes. Each optic artery splits into smaller vessels that ramify

3 20 Iain J. McGaw throughout the eye ~Fig. 1c!. A large lacuna on the dorsal surface of the eye ~sg! surrounds the sinus gland. A single artery continues from the sinus gland, terminating in the retina of the eye, where it splits into smaller capillary-like vessels. A number of other smaller vessels of approximately 100 mm diameter perfuse the medulla externa, medulla interna, and medulla terminata of the eyestalk. A smaller U-shaped vessel ~X! represents the arteries that surround the X organ within the eye ~Fig. 1c!. A thin membrane ~es! a few microns thick, was present on the ventral surface of the cast. This is part of the external eye sinus; after flowing through vessels of the eye, the hemolymph collects in this sinus, which is formed between the optic ganglia and the walls of the eyestalk. Anterolateral Arteries and Associated Branches The anterolateral arteries ~ALA! extend dorsally and anteriorly from the heart at approximately a 408 angle, relative to the anterior aorta ~Fig. 2a!. The main anterolateral artery serves as a source for a number of arteries. Two smaller arteries originate on the lateral surface: Close to the base of the artery a small branch exits at right angles to the main artery. This is the posterior gonadal artery ~PGA!. It supplies hemolymph to the posterior edge of the hepatopancreas and the gonads. A second larger branch, the gonadal artery ~GOA!, exits laterally at right angles to the main vessel about two-thirds along its length. Its course is superficial and it curves ventrally around the distal dorsal edge of the hepatopancreas and extends out to the lateral surfaces of the cephalothorax. The gonadal artery supplies hemolymph to hepatopancreas, as well as the gonads, and the hypodermis of the cephalothorax ~Fig. 2b!. Many fine branches originate along the length of the gonadal artery ~arrows indicate their point of origin; Fig. 2a!. These branch profusely over the superficial surface of the hepatopancreas and gonads. They supply hemolymph to the hypodermis of the ^ Figure 1. a: Corrosion cast of the anterior aorta complex ~AA! of Cancer productus. The end is enlarged, forming the cor frontale ~CF!. The small cerebral artery ~CA! arises anteriorly from the cor frontale, and the optic arteries ~OA! exit laterad to this structure. The optic arteries perfuse the eyestalks and smaller vessels, the lateral optic arteries ~LOA! branch to supply hemolymph to the musculature of the eyestalks. b: The cerebral artery ~CA!. The dense network of vessels ~on! supplies regions of the optic and oculomotor neuropiles of the supraesophageal ganglion, while the anterior branches of the cerebral artery perfuse the olfactory lobes ~ol!. c: Scanning electron micrograph ~ 25! of the left eyestalk. Blind-ending vessels from the optic artery ~OA! deliver hemolymph to the retina of the eye. An enlarged area of the dorsal surface of the eye represents the sinus gland ~sg! and vessels in the medial ventral area wrap around the X-organ complex ~X!. Part of the sinus ~ES! formed between the ventral surface of the eye and the carapace is still intact.

4 The Crustacean Circulatory System 21 Figure 2. The left anterolateral artery ~ALA! complex of Cancer irroratus. The cardiac stomach artery ~CSA! supplying hemolymph to the gonads branches off the inner surface of the anterolateral artery. At its end the anterolateral artery splits into the mandibular artery ~MA! and antennal gland artery ~AGA!, with the antennal gland ~ag! appearing as a dense mass. The gonadal artery ~GOA! exits at right angles on the outside of the anterolateral artery; this artery and its subbranch the posterior hepatic artery ~PHA! supply hemolymph to the gonads and hepatopancreas. The arrows indicate the point of origin of fine arteries arising along the length of the gonadal artery that were lost on the cast. b: These small arteries originating from the gonadal artery ~GOA! branch over the surface of the hepatopancreas, supplying hemolymph to this organ and the hypodermis of the carapace. c: They bifurcate along the anterior edge of the cephalothorax ~arrows!. The upper branch extends into each of the teeth along the anterior edge of the cephalothorax ~ct!. d:. The lower branches ~ventral view of crab! supply hemolymph to the hypodermis of the suborbital and subhepatic regions of the ventral carapace ~arrows!. anterior cephalothorax ~Fig. 2b!. Branches of these vessels ~ct! bifurcate terminally in at the anterior edge of the carapace ~arrow!. One branch extends into each of the teeth along the anterior margin of the carapace ~Fig. 2c!. The other branch ~arrows! continues perpendicularly and ventrally, supplying the hypodermis of the suborbital and subhepatic regions of the ventral carapace ~Fig. 2d!. A smaller lateral branch of the gonadal artery, the posterior hepatic artery ~PHA!, provides hemolymph to the posterior dorsal part of the hepatopancreas and carapace ~Fig. 2a!. The first medial branch of the main anterolateral artery ~Fig. 2a! is the cardiac stomach artery ~CSA!. This artery dips ventrally and anteriorly, passing through the gonads; additional branches leave the gonads and supply hemolymph to the pyloric and cardiac stomachs as well as the cardiopyloric muscles ~Fig. 2a!. The anterolateral artery continues its course toward the head region and splits into two vessels. The outside branch is the mandibular artery ~MA!, which supplies the external adductor muscles of the mandibles through a number of subbranches ~Fig. 2a!. The medial branch is the antennal gland artery ~AGA!. The antennal gland ~ag! lies below the main antennal gland artery and is served by the small coelomosac artery ~CCA!, which branches off the main artery ~Fig. 3a!. The antennal gland artery continues its course anteriorly and dips slightly, where three branches exit. Two of these branches continue their course anteriorly to supply the bladder, musculature of the antennae, and antennules and the eye retractor muscles ~Fig. 2a!. The third branch was lost on the preparation ~arrow!; its course is 1808 to the main vessel, flowing back toward the midline and following the course of the anterior aorta.

5 22 Iain J. McGaw Figure 3. a, b: Scanning electron micrograph ~dorsal view! of the antennal gland of Cancer magister at magnifications of 18 and 45, respectively. The gland is supplied by the coelomosac artery ~CCA!, which then divides into a plexus over the surface of the end sac ~esp!. This plexus continues into a highly convoluted series of vessels ~l! within the labyrinth of the antennal gland. These finally drain into a venous sinus ~vs! that covers the posterior surface of the gland. c: The area of the labyrinth indicated by a circle in a, is magnified 100, showing the convoluted network of vessels. d: Further enlargement ~ 300! reveals a series of circular vessels surrounding an open area ~arrow!. Scanning electron micrographs of the antennal gland of Cancer magister reveal its detail ~Fig. 3a!. The coelomosac artery ~CCA! immediately branches into a plexus of vessels ~esp!, over the dorsal surface of the end sac ~Fig. 3b!. This plexus gives rise to a convoluted network of capillaries ~l! that invade the tissues of the labyrinth of the antennal gland ~Fig. 3c,d!. These vessels are approximately mm in diameter. They form regular circular patterns that wrap around the columnar epithelial cells in the labyrinth. After flowing through the labyrinth capillaries, the hemolymph drains into a venous sinus ~vs; Fig. 3b! on the dorsal surface of the antennal gland. Hepatic Arteries The paired hepatic arteries ~HA! arise anteriorly and ventrally from the heart at about a 108 angle relative to the anterior aorta ~Fig. 4a!. ~The fine vessels have been removed on the corrosion cast to show the anatomy of the major vessels more clearly.! Almost immediately, these arteries dip ventrally under the stomach and laterally into the hepatopancreas, where three subbranches extend into the lobes of the hepatopancreas. Each hepatic artery gives rise to many capillary-like vessels that ramify profusely within the hepatopancreas ~Fig. 4b!. High magnification shows ~Fig. 4c e! a large number of terminally ending capillaries perfusing the hepatopancreas, forming a dense intertwining network of smaller vessels only mm in diameter ~Fig. 4e!. From here vessels connect into the lacunae~arrows! of the hepatic sinus ~Fig. 4e!. A lateral branch, the pyloric hepatic artery ~PYA!, exits each main hepatic artery soon after it leaves the heart. This vessel bifurcates almost immediately: One branch curves anteriorly downward and under the pyloric and cardiac stomachs, supplying hemolymph to these tissues. This vessel terminates close to the ventral thoracic

6 The Crustacean Circulatory System 23 Figure 4. a: Corrosion cast of the right hepatic artery ~HA! of Cancer anthonyi. The main branch divides within the hepatopancreas. Branching off the inside of the main artery, the pyloric hepatic artery ~PYA! flows anteriorly under the stomach and posteriorly along the midgut. b: Many fine vessels branch off the main artery ~HA! to form a dense network of capillaries ~hc!. c: Scanning electron micrograph ~ 18! of an area shows a fine network of vessels. d, e: Higher magnification of the outlined area ~ 35 and 85! reveals a series of blind-ending interdigitating vessels that extend to the borders of the hepatopancreas. artery between the third and second maxilliped vessels ~Fig. 6b!. The other branch issues posteriorly from the pyloric hepatic artery and runs along the length of the midgut ~Fig. 4a!. Posterior Aorta The posterior aorta ~PA; Fig. 5! is the smallest diameter vessel; it originates on the posterior ventral side of the heart. There are obvious differences in the anatomy of this vessel between male ~Fig. 5a! and female crabs ~Fig. 5d!. Two arteries branch from the main artery close to its origin with the heart; these are the posterior lateral arteries ~PLA!. Only one is preserved on each of the casts ~Fig. 5b,e!. These coil through the posterior midgut caecum, dipping ventrally and anteriorly around the end of the midgut. Two side branches exit the posterior lateral arteries, descending beneath the edges of the carapace and around the periphery of the posterior midgut cecum ~Fig. 5b,e!. These vessels join with the inferior abdominal artery in the region of the second abdominal segment. The posterior aorta itself descends through the abdomen and there are regular branches coming off the posterior aorta ~asa! supplying the tissues and muscles of each abdominal segment. These are larger in the female ~Fig. 5e! compared with the male ~Fig. 5b!, reflecting differences in the sizes of the abdomen ~Fig. 5a,d!. The posterior aorta continues along the hindgut, but soon after bifurcates ~hga!, traversing either side of the hindgut to the anus in the telson, and giving off smaller branches to the tissues of the rectum and abdomen ~Fig. 5b,e!. A transverse view of the posterior aorta shows further detail. In the male ~Fig. 5c!, paired arteries ~gpa! issue perpendicularly from branches of the fourth abdominal segment arteries. These supply hemolymph to the paired gonopods. In the female ~Fig. 5f! two pairs of arteries ~pla! issue perpendicularly from each abdominal segment artery ~asa!. These supply blood to each of the pleopods. Sternal Artery Exiting from the ventral surface of the heart close to the posterior aorta, the sternal artery ~SA! is the largest in

7 24 Iain J. McGaw Figure 5. The posterior aorta of Cancer magister exits posteriorly and ventrally from the heart and supplies the abdomen and associated structures. The abdomen of the male ~a! is narrower than that of the female ~d; nongravid individual!. This is reflected in the anatomy of the vessel ~b, e!. The main posterior aorta ~PA! gives off side branches to each abdominal segment ~asa!. The artery bifurcates terminally, supplying the hindgut region ~hga!. Paired posterior lateral arteries ~PLA! branch from the main artery, close to its origin from the heart; only one is preserved on each of the casts. Lateral views of the posterior aorta of the male ~c! showing vasculature of the paired gonopods ~gpa!. f: In the female each of the pleopods receives hemolymph from vessels ~pla! that branch off each abdominal segment artery. the circulatory system; its course is directly ventral ~Fig. 6a!, where it passes the midgut. In the present study, this artery passes on either the right or left side of the midgut. After the sternal artery passes the midgut, it turns 908 anteriorly and passes through the fused thoracic ganglia. It then turns 908 ventrally and it ends at a point in the thorax above the telson, where it bifurcates. The ventral portion of the sternal artery gives rise to pereiopod arteries 3 5 ~P3 P5! with the inferior abdominal artery issuing from the posterior end. The other end of the sternal artery proceeds anteriorly, giving rise to the first two pereiopod arteries ~P1 and P2! and the ventral thoracic artery ~VTA!. Inferior Abdominal Artery The inferior abdominal artery ~IAA! exits from the posterior end of the sternal artery ~Fig. 6a! and proceeds posteriorly and dorsally, following the course of the sternal artery. The inferior abdominal artery then turns anteriorly through 1808 along the thorax. Its course appears to be superficial over the surface of the thorax, giving off smaller side branches into the tissues of the thoracic sternites. Two smaller side branches issue from the main artery and pass through the abdomen. They join with the posterior lateral arteries in the region of the second abdominal segment, supplying hemolymph to the hindgut and muscles of the abdomen. Finer

8 The Crustacean Circulatory System 25 Figure 6. a: The sternal artery ~STA! of Cancer magister exits ventrally from the heart. Five pairs of arteries ~P1 P5! exit laterally from the main artery, supplying hemolymph to the pereiopods. The inferior abdominal artery ~IAA! exits from the posterior surface of the sternal artery. b: The ventral thoracic artery ~VTA! exits from the anterior end of the sternal artery, supplying the mouthparts. Three paired branches arise from the ventral thoracic artery. The first two branches ~mx2, mx3! supply hemolymph to the exopodite ~ex2, ex3! and endopodite ~en2, en3! of the second and third maxillipeds. A dense network of vessels ~scp! branches from the outside of the anterior pair of arteries. These perfuse the muscles of the scaphognathite. A smaller branch arises from the left scaphognathite vessel and splits to supply the first maxilliped ~mx1! and the mandibles ~mn!. c: Photograph of mouthparts of C. magister. The third maxilliped has been removed on one side to expose the underlying structures. The position of the scaphognathite ~scp! is also indicated. ^ vessels also radiate from the main inferior abdominal artery and supply the distal portion of the midgut. Ventral Thoracic Artery The ventral thoracic artery ~VTA! issues anteriorly from the sternal artery ~Fig. 6a! and is broadest at its origin with the sternal artery. This artery and its branches supply the mouthparts of the crab ~Fig. 6c!. The ventral thoracic artery branches into seven vessels: three paired arteries that exit laterally, and a smaller single artery that exits anteriorly, which gives rise to yet two more pairs of arteries ~Fig. 6b!. The first paired arteries ~mx3! exit at about 758 relative to the main vessel and supply the third maxillipeds. The inner branch curves into the endopodite ~en3! and the outer branch extends into the exopodite of the third maxilliped ~ex3!. A second pair of vessels ~mx2! arises from the ventral thoracic artery, anteriorly to the first pair; these supply hemolymph to the second maxilliped ~mx2!. In the cast shown, the artery on the right side originates anteriorly to the branch on the left side. These arteries bifurcate, continuing their course anteriorly. The outer branch supplies the exopodite and meropodite of the second maxilliped ~ex2!; whereas the inside branch supplies the endopodite of the second maxilliped ~en2!. At the anterior end of the ventral thoracic artery is a final pair of arteries ~scp!, which are greater in diameter than the first two pairs. These arteries exit anteriorly and laterally at an approximate angle of 458 to the main ventral thoracic artery. They split into three smaller vessels that give rise to a dense network of capillarylike vessels supplying the scaphognathite muscles. A single vessel traverses the length of the scaphognathite, although this was lost on the current preparation. A final branch of the ventral thoracic artery arises on the anterior surface of the left scaphognathite artery ~scp!. Almost immediately this vessel bifurcates, and within a short distance the vessel bifurcates again. The outer branches supply hemolymph to the first maxillipeds ~mx1! and the inner branches supply the mandibles ~mn!. Scanning electron micrographs show the detail of some of the arteries of the ventral thoracic system more clearly. The scaphognathite muscles are highly vascularized ~Fig. 7a,b!. Small vessels of approximately 20 mm diameter terminate in the muscles of the scaphognathite ~Fig. 6b!.

9 26 Iain J. McGaw

10 The Crustacean Circulatory System 27 Figure 7. a: Scanning electron micrograph ~ 18! shows the complex vasculature of the scaphognathite ~scp!. b: Enlargement ~ 55! of an area showing the fine blind-ending vessels ramifying throughout the scaphognathite muscle. c: Scanning electron micrographs ~ 18! of the vessel supplying the exopodite ~ex3! of the third maxilliped. d: Many fine vessels branch from the main vessel ~ 43!. e, f: Scanning electron micrograph of the exopodite of the third maxilliped showing the small vessels draining into a series of lacunae ~exs!. These then empty into a sinus ~arrow! formed between the musculature of the exopodite and the carapace. ^ The small vessels supplying the exopodites of the third maxillipeds are also complex ~Fig 7c!. These vessels are approximately mm in diameter. At higher magnification ~Fig. 7d!, a series of blind ending capillary-like vessels are evident; some of the finer vessels are only 10 mm in diameter. These supply hemolymph to the tissues of the exopodites. From here the hemolymph collects in lacunae ~exs! between the tissues ~Fig. 7f!. Hemolymph from these lacunae then drains into a sinus ~Fig. 7e! formed between the tissues of the exopodite and the carapace wall; this connects with the anterior portion of the ventral thoracic sinus. Pereiopod Arteries The first paired branches ~P1! of the anterior portion of the sternal artery exit laterally and anteriorly, supplying hemolymph to the chelae ~Fig. 8a!. This pereiopod vessel traverses the meropodite ~m!, and carpopodite ~c!, giving off small branches at regular intervals that divide among the muscles. Three branches ~p! supply hemolymph to the posterior extensor and anterior flexor muscles within the propodite. The main vessel ~P1! bifurcates at the dactylopodite ~d! with each vessel extending to the tips of the chelae ~Fig. 8b!. Within the chelae smaller blind-ending branches extend out to the edge of the carapace, supplying the musculature of the chelae ~Fig. 8b,c!. Hemolymph from these vessels collects in lacunae, forming a dense network within the dactylopodites ~Fig. 8d!. These lacunae ~p1s! and smaller blind-ending vessels are evident in the scanning electron micrographs ~Fig. 8e,f!. The lacunae eventually drain into a sinus formed in between the musculature and the edge of the carapace ~not shown!. The second set of paired arteries ~P2! exit from the anterior portion of the sternal artery, posterior to the first pair ~P1!. They also flow laterally and anteriorly supplying hemolymph to the second pair of pereiopods ~Fig. 6a!. The other three pairs of arteries arise from the posterior end of the sternal artery. The third pair of arteries ~P3! exit laterally and posteriorly, providing hemolymph to the third pair of pereiopods. The fourth pereiopod arteries follow a similar plan to the third pereiopod arteries. However the fifth pereiopod arteries do not arise directly from the sternal artery. Instead they branch off pereiopod artery 4, a short distance after it exits from the sternal artery. Their course is lateral and posterior and they supply hemolymph to the fifth pair of pereiopods. On each pereiopod artery, a smaller branch arises on the superior surface; this is the branchial artery ~not shown!. This artery wraps around the base of the gills. From here the vessel turns through 1808 under the gill somites, supplying hemolymph to the tissues of these structures. Pereiopod arteries 2 5 are anatomically similar. Many fine capillary-like vessels arise from each artery, the density of which is greatest within the muscles of the thoracic sterna; this area is highly vascularized ~Fig. 6a!. Each pereiopod artery extends into the tip of the dactylopodite and capillary-like vessels branch extensively along the entire length, supplying hemolymph to the muscles of the pereiopods ~Fig. 9a!. From here blood collects in a sinus ~LS! formed between the muscles of the pereiopods and the carapace wall. Hemolymph flows back along the leg sinus where it joins with the infrabranchial sinuses. Each leg sinus is less than 10 mm thick and is covered in a series of pores ~Fig. 9b, arrows!. Scanning electron micrographs ~Fig. 9c! show these structures consist of a collar about 30 mm in diameter with a small short vessel of approximately 5 mm diameter exiting from the center. This may represent vascular supply to the sensory setae covering the outer surface of the integument. Sinuses Hemolymph collects in sinuses before flowing over the gills and back to the heart. Most of the sinuses are discrete channels rather than arbitrary spaces between organs, and their form is determined by the boundaries of specific organs or muscles. However, because of their delicate form, they were difficult to demonstrate on the corrosion casts. Spent hemolymph from the anterolateral arteries and anterior aorta drains into a dorsal sinus ~DS!. Intact dorsal sinuses were difficult to demonstrate, because under abnormally high pressures such as those generated during infusion of Batsons Monomer, the sinuses may be blown apart, thus exaggerating their form and interpretation. The lacunae of the dorsal sinus appear as a fine network of vessels covering the anterior surface of C. antennarius ~Fig. 10a!. The cast below ~Fig. 10b! shows a more typical preparation; the right side of the dorsal sinus has blown apart, filling the space between the stomach and superficial surface of the hepatopancreas. On the left side the relative positions of hepatic capillaries, anterolateral artery, and the anterior aorta can be clearly seen. Hemolymph returning from the eyestalks and the anterior portion of the crab collects in the eye sinus. In the

11 28 Iain J. McGaw Figure 8. a: Corrosion cast of the first pereiopod artery of Cancer antennarius showing the meropodite ~m!, carpopodite ~c!, propodite ~p!, and dactylopodite ~d! regions of the chelae that it serves. b: Pereiopod artery 1 ~P1! bifurcates terminally in the dactylopodites of the chelae. c: Enlargement of the end of the chelae showing the fine vasculature. d: Corrosion cast showing the sinus system of a chelae. e, f: Scanning electron micrographs of the sinus of the chelae. Most of the fine vessels are connected via a series of tubular lacunae ~p1s!; others terminate as blind ending vessels ~arrows!.

12 The Crustacean Circulatory System 29 preparation shown ~Fig. 10c!, the anteriorly directed vessels and stomach have been removed. The eye sinuses extend around the anterior edge of the stomach. From here they continue posteriorly and ventrally, where they join with branches of the dorsal sinus. The boundaries of the eye sinus are determined by the stomach and associated muscles and the edge of the hepatopancreas. The eye sinus joins with sinuses from the mouthparts and the ventral thoracic sinus ~VTS! in the ventral region of the crab ~Fig. 10c!. The hepatic sinus ~HS! forms a thin covering over the surface of the hepatopancreas and the hepatic capillaries ~hc; Fig. 10d!. On most of the corrosion casts the hepatic sinus was only partially preserved as a series of lacunae covering the anterior surface of the hepatopancreas. Removal of the hepatic capillary mass ~Fig. 10e! shows the sinus invades the hepatopancreas via vertical channels ~arrows!. A large anterior vein ~AV! was present in preparations ~Fig. 10d!. This joins with the hepatic sinus and part of the dorsal sinus. From here it flows along the anterior edge of the carapace before dipping ventrally, where it joins with the eye sinus ~ES! in the head region of the crab. Hemolymph from the dorsal, eye, and hepatic sinuses collects in a large ventral thoracic sinus on the ventral side of the crab ~VTS!. The ventral thoracic sinus radiates out between the muscles of the thoracic sterna as the branchial sinuses ~BrS; Fig. 10f!. The branchial sinuses form distinct lacunae between the muscles of the legs and the carapace. The branchial sinuses then fuse with the infrabranchial sinus ~IBS! that runs along the posterior edge of the gills. From here the hemolymph flows over the gills, where it is oxygenated. Branchiostegal Circulation A complex series of sinuses and vessels make up the branchiostegal circulation, which acts as a venous return route that bypasses the gills. These sinuses and vessels cover the ventral surface and part of the dorsal surface of the branchial chambers ~Fig. 11a,c!. The branchiostegal sinus ~BS! originates on the ventral side of the branchial chamber where the pulmonary vein ~PV! branches from the junction of the eye and dorsal sinus ~Fig. 11c!. A fine series of vessels and lacunae cover the ventral floor of the branchial chamber ~Fig. 11d!. These lacunae are directed posteriorly, flowing up and over the posterior edge of the branchial chamber ~Fig. 11b!. Blood is returned to the pericardial sinus ~Fig. 11b! via the marginal ~mv! and lateral veins ~lv; Fig. 11b!. Figure 9. a: Corrosion cast of the fifth pereiopod artery of Cancer antennarius ~P5!. Blood eventually collects in a sinus ~LS! that surrounds each artery, much like a glove. b: Scanning electron micrograph ~ 130! of the outer surface of the leg sinus; arrows indicate a series of pores. c: Higher magnification ~ 550! reveals a cylindrical structure with a small vessel approximately 50 mm in length and 5 mm in diameter. These supply hemolymph to each of the hairlike setae. Gill Vessels Hemolymph collects in the infrabranchial sinus ~IBS! at the base of the gill ~Fig. 12a!. From here it flows along the afferent branchial vein ~ABV! and through the gill lamellae ~GL!, which lie close to each other ~Fig. 12b!. Individual lamellae are held apart with spacing nodules ~sn! on the anterior edge ~Fig. 12c!. Scanning electron micrographs show the detail of an individual lamella. Hemolymph flows

13 30 Iain J. McGaw Figure 10. a: The dorsal sinus ~DS! of Cancer antennarius showing the lacunae of the sinus. b: Corrosion cast of Cancer gracilis, showing the major anteriorly directed vessels: anterolateral ~ALA!, anterior aorta ~AA! and the mass of hepatic capillaries ~hc!. In this case the dorsal sinus ~DS! has been over-expanded during infusion of Batsons monomer c: Dorsal view of the eye sinuses ~ES! of Cancer productus. The sinuses collect hemolymph from the eyes; they continue their course posteriorly and ventrally around the edges of the stomach. The ventral thoracic sinus is evident on the ventral surface of the crab. d: Transverse view along the left anterior margin of the cephalothorax of Cancer productus. The mass of hepatic capillaries ~hc! is evident in the center. This is bounded on both side by the hepatic sinus ~HS!. A large anterior vein ~AV! exits anteriorly from the hepatic sinus; its course is ventral where it joins with the eye sinus. e: Removal of the hepatic capillaries exposes a series of vertically directed sinuses ~arrows!; these collect hemolymph from the hepatopancreas. f: Ventral view of a corrosion cast of Cancer gracilis. The ventral thoracic sinus ~VTS! is visible toward the center of the crab. This sinus gives rise to the branchial sinuses ~BrS!, which consist of a series of tightly fitting lacunae. The branchial sinuses connect with the infrabranchial sinus ~IBS!. From here the hemolymph flows through the gills. through the marginal canal ~mc! along the anterior edge of the lamellae. It then flows through many fine lamellar capillaries that cover the surface of the lamellae ~Fig. 12d!. The arrows indicate the direction of blood flow ~Fig. 12e!. After passing through the lamellae, hemolymph flows along the efferent branchial vein ~EBV! and continues into the branchiocardiac veins ~BCV; Fig. 12a!. From here, it drains into the pericardial sinus and back into the heart through six pairs of ostia. DISCUSSION During the past 40 years, vascular corrosion casting techniques have been used to study the microvasculature in an array of organs and tissues ~see Hossler & Douglas, 2001!. Such methodology produces an accurate three-dimensional representation of vessel structure and anatomy ~Kratky et al., 1989; Hossler & Douglas, 2001!.

14 The Crustacean Circulatory System 31 Figure 11. a: Dorsal view of a corrosion cast of Cancer anthonyi. The heart ~H! and ostia ~ost! are in the center. The branchiostegal sinus ~BS! covers much of the dorsal posterior surface of the animal. b: Enlargement of the posterior region of the branchiostegal sinus ~dorsal view! showing the marginal ~mv! and lateral veins ~lv!, which return hemolymph to the pericardial sinus. c: Ventral view of C. anthonyi showing the branchiostegal sinus ~BS! extending over the ventral surface of the branchial chamber. The sinus is supplied with hemolymph via the pulmonary vein ~PV!. The anterior vein ~AV! can be clearly seen on the surface of the hepatic capillaries. d: Scanning electron micrograph ~ 18! of part of the ventral branchiostegal sinus, showing the network of interdigitating lacunae that connect into the pulmonary vein ~PV!. Although the anterior aorta was the third largest artery diameter-wise, hemolymph flow rates through this artery are low. Less than 1% of the cardiac output is pumped through the anterior aorta of C. magister ~Airriess & McMahon, 1994, 1996; McGaw et al., 1994a, 1994b!. The cor frontale lies at the end of the anterior aorta ~Fig. 1!; it consists of tendons and two strips of striated muscle ~Steinacker, 1978!. Its exact function is unclear. It is thought to act as an auxiliary heart, aiding hemolymph flow into the supraesophageal ganglion ~Steinacker, 1978; Davidson & Taylor, 1995!; it may also behave as a variable elastic buffer by dampening pressure pulses to the cerebral vessels ~Davidson & Taylor, 1995!. Exiting anteriorly from the cor frontale, the cerebral artery supplies the supraesophageal ganglion ~Fig. 1b!. The cerebral artery ramifies throughout the optic and oculomotor neuropiles, and finer vessels continue around the olfactory lobe. In the green crab, Carcinus maenas, these vessels form a true closed system with both afferent and efferent capillaries perfusing the brain ~Sandeman, 1967; Abbott, 1971!. In the current preparation the presence of a true capillary network was inconclusive. It is possible that the infusion pressure was not great enough to force the viscous resin through the high resistance capillary network ~Abbott, 1971!. These delicate vessels may also have been lost during the tissue maceration process ~Kratky et al., 1989!. The paired optic arteries also branch from the cor frontale and supply hemolymph to the eyes and associated

15 32 Iain J. McGaw Figure 12. a: Corrosion cast of an excised gill of Cancer magister. Hemolymph flows from the infrabranchial sinus ~IBS! along the afferent branchial vein ~ABV! over the gill lamellae ~GL!, where it is oxygenated and into the efferent branchial vein. From here it flows back into the pericardial sinus via the branchiocardiac vein ~BCV!. b: Scanning electron micrograph ~ 25!, showing the afferent branchial vein and gill lamella. c: Scanning electron micrograph ~ 200! showing enlarged regions that are the spacing nodules ~sn! on each gill lamellae. d, e: Scanning electron micrographs ~ 18, 40! of an excised lamellae. Hemolymph flows through the marginal canal ~MC! and through a dense network of lamellar capillaries ~LC!. Arrows indicate the direction of blood flow.

16 The Crustacean Circulatory System 33 structures. In the green crab, C. maenas, the eyes receive an extensive network of vessels, some with afferent and efferent connections ~Sandeman, 1967!. Complete capillary networks were not evident in the present study. Sandeman ~1967! also had limited success in demonstrating these capillary networks repeatedly, when using ink injection. The eyes contain enlarged lacunae around the position of the sinus gland and X-organ complex ~Fig. 1c!. These organs are connected with hormone production and require a good blood supply to ensure effective hormone delivery to other areas of the body. After perfusing the eye structures, blood collects in an outer sinus. The eye sinuses were apparent in some casts as a very fine veneer ~Fig. 1c!, their form being determined between the retina and the outer wall of the eye. However, during excess perfusion pressure, the delicate eye sinuses appear as enlarged masses, similar to those reported by Davidson and Taylor ~1995!. The anatomy of the anterolateral arteries was more complex than previously described for cancrid crabs ~Pearson, 1908; Airriess, 1994!. These paired vessels split into several smaller branches that supply hemolymph to the hypodermis of the cephalothorax, the dorsal surface of the hepatopancreas, the gonads, mouthparts, and antennal glands. Blood flow into the anterolateral artery is controlled via contraction or relaxation of cardioarterial valves at the base of the artery ~Alexandrowicz, 1932!. However, each of the vessels that branch from the main anterolateral artery ~Fig. 2a! supply distinct organs that perform quite different physiological functions. Thus each of the branches of the main artery may not need to be perfused at the same rate. It therefore follows that hemolymph to the specific areas may be controlled separately. In a number of crustaceans ~Sicyonia ingentis, Homarus americanus, Panulirus interruptus! some of the major arteries contain layers of striated muscle that may help in redistribution of blood ~Burnett, 1984; Martin et al., 1989; Wilkens et al., 1997a!. There is no evidence of striated muscle in the anterolateral arteries of Cancer species ~Shadwick et al., 1990!. Nevertheless, arterial resistance ~and hence relative rate of blood flow! can be controlled by a variety of pericardial neurohormones ~Wilkens & Taylor, 2003!. Such mechanisms may allow differential rates of hemolymph flow through each of the subbranches of the anterolateral artery. In the cancrid crabs the antennal gland was well defined. However, it is not supplied directly by the antennal gland artery. Instead a smaller artery, the coelomosac artery, branches from the ventral side of the antennal gland artery. This splits into smaller capillaries that ramify within the labyrinth of the antennal gland. This narrowing of the artery probably allows a higher pressure for filtration within the labyrinth of the antennal gland. Thus, pressures in the main artery may not reflect what is happening downstream. The complexity of the Cancer antennal gland is similar to that reported for the paddle crab Ovalipes catharus ~Davidson & Taylor, 1995!, the crayfish Procambarus blandingi ~Peterson & Loizzi, 1974!, and the lobster H. americanus ~Miyawaki & Ukeshima, 1967!. In the blue crab, the blood supply to the antennal gland is not as complex ~Johnson, 1980; McGaw & Reiber, 2002!. This is unexpected because the blue crab is classed as a efficient osmoregulator ~Tan & Van Engel, 1966!, whereas the Cancridae are weaker osmoregulators ~Hunter & Rudy, 1975!. The paired hepatic arteries exit from the anterior and ventral surface of the heart and branch profusely within the hepatopancreas ~Fig. 4a!. These arteries are depicted as very small vessels in portunid crabs ~McLaughlin, 1983! and crayfish ~Reiber et al., 1997!. In C. magister these arteries have the second largest lumens and they receive 25 30% of the total cardiac output ~McGaw & McMahon, 1995, 1999!. They branch profusely within the hepatopancreas, terminating as blind-ending vessels. The diameter of some of the smaller vessels was approximately mm ~Fig. 4e!, which corresponds closely with those in the blue crab, C. sapidus ~Johnson, 1980! and the lobster H. americanus ~Factor & Naar, 1990!. The hepatic capillaries formed the densest network of vessels within the animal ~Fig. 4c e!. This was expected because the hepatopancreas must support all metabolic activities involved with digestion, including delivery of enzymes, transport of digested nutrients, and removal of digestive wastes ~Icely & Nott, 1992!. The blind-ending vessels connect to a well-defined hepatic sinus that forms a network of lacunae over the surface of the hepatopancreas ~Fig. 10d! and extends between the lobes of the hepatopancreas ~Fig. 10e!. In brachyuran crustaceans, the posterior aorta is the smallest diameter artery ~Pearson, 1908; Pyle & Cronin, 1950!. Estimates from corrosion casts show that it carries less than 2% of the blood supply and receives only 2 3% of total cardiac output ~McGaw & McMahon, 1995, 1999!. The posterior aorta, posterior lateral arteries, and inferior abdominal artery anastomose in the region of the second abdominal segment ~Fig. 5!. This is somewhat different from that shown by McLaughlin ~1983!, where the inferior abdominal artery joins the posterior aorta in the telson. The co-lateral supply of hemolymph to the abdomen and hindgut by the posterior aorta, posterior lateral arteries, and inferior abdominal artery may help maintain hemolymph flow to these tissues when flow in either artery becomes occluded by movement of the abdomen or the presence of food in the midgut or hindgut. Even the reproductive structures were vascularized by a series of small-diameter vessels ~Fig. 5c,f!. Such an intricate perfusion has not been reported before in Cancer crabs ~Pearson, 1908; Airriess, 1994!. The sternal artery is the largest artery in the system. This artery and its associated branches can receive up to 80% of the total cardiac output during periods of increased activity ~DeWachter & McMahon, 1996; McGaw & McMahon, 1998; I.J. McGaw, in press!. The sternal artery is valved at its origin, but control of blood flow into the peripheral regions served by this arterial complex is probably influenced by changes in downstream resistance, the possible result of skeletal muscle contractions ~Wilkens, 1997; Wil-

17 34 Iain J. McGaw kens et al., 1997a,b! or neurohormonal control ~Wilkens & Taylor, 2003!. Each pereiopod and its associated muscles were perfused through many fine capillary-like vessels. The promotor, remotor, levator, and depressor muscles of the thoracic sterna were highly vascularized by smaller vessels radiating off of the main pereiopod arteries ~Fig. 6a!. Such an intricate network of vessels has not been reported for Cancer spp ~Pearson, 1908; Airriess, 1994!. The muscles of the chelae were supplied by pereiopod artery 1 ~Fig. 8b,c!, with branches reaching into the tip of each claw. This perfusion is more complex than in the blue crab, C. sapidus. Cancer crabs are able to exert a higher and more continuous force compared with blue crabs ~Taylor, 2000! and therefore require a greater hemolymph demand to supply oxygen to the muscle. In fact, each individual muscle fiber may be perfused by as many as seven or eight tiny vessels ~Govind & Guchardi, 1986!. Once hemolymph has flowed through the chelae it collects in well-defined lacunae ~Fig. 8e,f!. The lacunae drain into larger sinuses, which are discrete units, formed between the outer muscle layers and the carapace. This sinus is well defined in pereiopod arteries 2 5 ~Fig. 9a!. Smaller peglike vessels exit on the outer surface of this sinus. These connect to the setae on the legs, possibly the hair peg organs, which have a lymph space surrounding them ~Schimdt, 1989!. It is the presence of sinuses, rather than a complete series of veins, that defines the circulatory system as open. However, there is disagreement in the literature about how well defined these sinuses are. In the blue crab, C. sapidus ~Pyle & Cronin, 1950! and the edible crab Cancer pagurus ~Pearson, 1908! many of the sinuses are reported to be so ill defined that they defy successful demonstration. Nevertheless, over a century ago Haeckel ~1857! proposed that no unbounded lacunae exist in the crustacean system. Major sinuses are bordered by fibrous connective tissue and the lacunae by basal lamina directly on the organ that they bathe ~Johnson, 1980!. The distinction between sinus and capillary then becomes less distinct, suggesting a more organized structure. In the present study, most of the sinuses appeared as a fine layer of lacunae; however, because of their delicate form, some were lost on the corrosion casts while the tissue was being macerated, and during subsequent handling of the casts. Thus, this is one of the limitations of the corrosion casting methodology. Nevertheless the present study suggests that the sinuses are discrete channels rather than arbitrary spaces between organs; their form is determined by the boundaries of specific organs or muscles. During abnormally high pressures such as those generated during infusion of Batsons Monomer, the sinuses may be blown apart, thus exaggerating their form and interpretation ~Fig. 10a,b!. The fact that all areas of the arterial system were perfused in the present study, including the return channels, the gills, and branchiostegal sinuses, suggests that casts were complete. Thus the representation of the sinuses is probably accurate. Once hemolymph has bathed the tissues it collects in the ventral thoracic sinus below the heart. The ventral thoracic sinus radiates out between the muscles of the thoracic sterna as the branchial sinuses, which fuse with the infrabranchial sinus that runs along the posterior edge of the gills ~Pearson, 1908!. From here, hemolymph flows through the gill lamellae via the afferent and efferent branchial veins, which are distinguished from sinuses by having a definite shape and function as return channels for blood ~Maynard, 1960!. Hemolymph moves through this extensive network of channels and/or passes along the marginal canal of the lamellae, which provide an extensive surface area of exchange with inflowing water ~Fig. 12d,e!. Once hemolymph has passed through the lamellae it drains into the efferent branchial vein and continues into the branchiocardiac veins, which carry arterial blood back to the heart. An alternative, dorsal venous return route via the branchiostegal circulation has been proposed ~Taylor&Greenaway, 1984; Farrelly & Greenaway, 1987; Greenaway & Farrelly, 1990; Taylor & Taylor, 1992!. The branchiostegal circulation appears as a fine mesh-work of lacunae that covers the posterior dorsal surface of the branchial chamber and extends around the pericardial cavity ~Fig. 11!. Itis supplied with hemolymph from the dorsal and eye sinuses. Hemolymph returns to the pericardial cavity by way of lateral collecting vessels ~Fig. 11b!. The branchiostegal circulation is well developed in terrestrial crabs ~Taylor&Greenaway, 1979! and amphibious species ~Greenaway & Farrelly, 1990; Greenaway et al., 1996!; in these animals it is used for aerial gas exchange. In Cancer species it was less well developed. Because members of this family are essentially subtidal, its exact role is unclear, but may be used for oxygen extraction during periods of reversed ventilation ~Davidson & Taylor, 1995!. Although the branchiostegal sinus can bypass the gills, the partitioning of venous return between the branchial and branchiostegal circulations has not yet been measured ~Taylor & Taylor, 1992!. The anatomy of the circulatory systems of the different species from the family Cancridae was very similar, but was more complex than previously described ~Pearson, 1908; Airriess, 1994!. All areas of the body were highly perfused by arteries, arterioles, and fine capillary-like vessels. The presence of sinuses, rather than a complete venous return system, defines the circulatory system as open. However, even the sinuses were discrete units, most of which branched into well-defined lacunae. In line with the anatomical intricacy, physiological control is also complex, with pressures that rival some simple vertebrate systems ~McMahon & Burnett, 1990!, and neurohormonal control mechanisms that allow fine tuning of cardiac function and regional hemolymph flow ~reviewed in McGaw & McMahon, 1999; Wilkens, 1999!. The present study complements and extends recent work on the cardiovascular system of the blue crab ~McGaw & Reiber, 2002!, showing that in Cancridae the circulatory system is even more complex. This supports