ANIMAL PHYSIOLOGY

Hydraulic control of tuna fins:A role for the lymphatic system invertebrate locomotionVadim Pavlov,1*† Benyamin Rosental,1,2* Nathaniel F. Hansen,1 Jody M. Beers,1

George Parish,3 Ian Rowbotham,3 Barbara A. Block1†

The lymphatic system in teleost fish has genetic and developmental origins similar tothose of the mammalian lymphatic system, which is involved in immune response and fluidhomeostasis. Here, we show that the lymphatic system of tunas functions in swimminghydrodynamics. Specifically, a musculo-vascular complex, consisting of fin muscles, bones,and lymphatic vessels, is involved in the hydraulic control of median fins. This specializationof the lymphatic system is associated with fish in the family Scombridae and may haveevolved in response to the demand for swimming and maneuvering control in thesehigh-performance species.

The lymphatic system of vertebrate animalsplays a key role in osmoregulation and im-mune response. Although developmentaland molecular markers indicate genetic or-igins similar to those in mammals (1, 2), the

lymphatic system in teleost fish, the most diverseclass of vertebrates, might perform additionalfunctions (3–8). A variety of experiments havedescribed the lymphatic system in teleost fish,illustrating that it is a separate, parallel circula-tory system (1, 5, 7, 9, 10). Current knowledge of

the lymphatic system in fish assumes that itspresence and distribution do not appear to beexclusively linked to phylogenetic position, butrather to the physiological adaptations of thespecies (4, 6). The lymphatic system has beenhypothesized to be more extensive in tunas thanin other teleost fish (3, 4), owing to their elevatedblood pressures (11), high metabolic rates in somespecies (12, 13), and high gill surface areas (14) thatlikely increase osmoregulatory demands (12, 15).However, the lack of data on the morphology of

the lymphatic system in tunas makes it difficultto determine its function. Here, we use a multi-disciplinary approach, applying vascular injection,as well as histological, cytological, and immuno-logical methods, video observations of swimming,computer-aided design (CAD), and computer fluiddynamics, to study the role of the lymphatic sys-tem of Pacific bluefin (Thunnus orientalis) andyellowfin tunas (Thunnus albacares).Dissections of tuna fin musculature and vascu-

lature revealed a large vascular sinus (VS) locatedat the base of both the second dorsal and analfins (Fig. 1A). Perfusion of a casting compound(Microfil) into the VS exposed a musculo-vascularcomplex not previously described in morpho-logical descriptions of tuna fins (Fig. 1 and figs.S1 and S2). Morphologically, the sinus creates achamber formed by the bone and cartilage tissues.The dome of the chamber is formed by the prox-imal parts of the fin rays, whereas the ventralsurface is formed by the adjacent proximal partsof pterygiophores supporting the fin (Fig. 1A).The junction between the fin rays and pterygio-phores restricts lateral movements of the fin rays.The VS is connected with vessels that form a vas-cular network around the fin and channels located

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1Department of Biology, Stanford University, Hopkins MarineStation, 120 Oceanview Boulevard, Pacific Grove, CA 93950,USA. 2Institute for Stem Cell Biology and RegenerativeMedicine, Stanford University School of Medicine, Stanford,CA 94305, USA. 3Tuna Research and Conservation Center,Monterey Bay Aquarium, Monterey, CA 93950, USA.*These authors contributed equally to this work.†Corresponding author. Email: bblock@stanford.edu (B.A.B.);pavlov.v.v@gmail.com (V.P.)

Fig. 1. Morphology of the musculo-vascularsystem in Pacific bluefin tuna median fins.(A) Cross section of the second dorsal fin (SDF)sinus.Top left inset: A dissecting scope image ofthe SDFcross section. Bar, 1 mm. Abbreviations: finrays (FR), expandable vascular channels (VC),incompressible chamber of the vascular sinus (VS),scales (SC), inclinatormuscles (IM) surround lateralvessels connected with VS, epaxial musculature(EPM), proximal parts of pterygiophores (PT),erector muscles (ERM), and depressor muscles(DM). (B) Cross section of the depressed (left) anderected (right) SDF shows collapsed and expandedVC between fin rays. (C) Three-dimensionalstructure of the musculo-vascular system in SDF.PTand some FR have been removed for illustrativepurposes. FR, connective tissue (CT) sheathbetween skin and IM, EPM, and compressiblevascular channels (CVC) continue around the firstdorsal fin (FDF). (D) Cross section of the FDF. FR ofthe folded FDF in the groove on the fish’s back, CVCbeneath the SC layer, IM, ERM and DM, and EPM.(E) Blue Microfil injection into the incompressiblechamber of theVS at the base of the SDF; skin withthe scales has been removed. Bar, 2 cm. FR of theerected FDF,CVC around the FDF, and connection ofthe CVC with the VS incompressible chamberat the base of the SDF; blue Microfil compoundinjected into the sinus reveals the expanded vascularchannels between FR of the SDF.

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between the fin rays (Fig. 1, B and C). The vascu-lar network of the second dorsal fin is connectedwith large vascular channels around the firstdorsal fin (Fig. 1, C to E).This common structure of the musculo-vascular

complex was observed in both second dorsal andanal fins: The inclinator muscles contact the soft-

walled vessels that are connected with the VSthat extends into the expandable channels be-tween the fin rays (Fig. 1, A and C, and figs. S1to S3). The second dorsal and anal fin erectionis hypothesized to be under the control of themusculo-vascular complex, including involvementof the erector, depressor, and inclinator muscles,

as well as vascular vessels. We hypothesize thatcontraction of inclinator muscles causes inflowof fluid from the soft-walled vessels to the in-compressible fin sinus and then to the expand-able fin channels (Fig. 1B). Increase of pressurein the fin channels facilitates the effort of theerector muscles and counteracts the work of thedepressor muscles, thus providing fine adjust-ments of the fin erection (Fig. 2, A and B, andmovie S1). To test this hypothesis, we pumpedsaline solution into the fin sinus of a recentlydeceased tuna to simulate the contraction ofthe inclinator muscles and alter pressures in finchannels. Increased pressure in the range of 17to 48 kPa caused fin erection and decreased finsweep angle (L) by up to 14° (movie S2 and fig. S4).The pressure-driven fin erection in the absenceof erector muscle effort is evidence of the hy-draulic effect in the fins, although the contribu-tion of specific muscles in fin movement remainsunclear.To understand how the pressure-driven fin

erection corresponds to the range of fin move-ments and behavior of living fish, we recordedand analyzed videos of swimming tunas in a cap-tive facility. Video data were categorized by thebehavior patterns of bluefin and yellowfin tunasbefore, during, and after feeding periods and bymeasured sweep angle of the second dorsal fin(fig. S4) that moves synchronously with the analfin. The fin erection depended on swimming be-havior and varied from L = 73.9 ± 4.3° (mean ±SD) during rectilinear movement when no feedingoccurred to L = 60.3 ± 7.2° (mean ± SD) duringfrequent changes of movement direction duringfeeding times [P < 0.001, repeated measures anal-ysis of variance (ANOVA); fig. S5 and table S1].Additional analyses of free-swimming tunas at alarge exhibit tank in the Monterey Bay Aquar-ium revealed that the maximal second dorsal finerection was significantly associated with turningmaneuvers (table S2). Together, these data suggestthat median fin erection increases from cruisingbehavior with predominant rectilinear movementto searching and feeding behavior with frequentchanges of movement direction. The results alsoindicate that the pressure-driven fin erection inthe in vitro experiment (movie S2) falls within thenatural range of fin erection in swimming tuna.Median fins in fishes are typically oriented in

the vertical plane and involved in control of pos-ture and swimming trajectories (16). Median finsof tunas are analogous to thin, symmetrical hy-drofoils generating sideways lift force (17) whenthe fin plane makes an angle to the fluid flowdirection (Fig. 2, C to E). Movable fins, capable ofchanging their area and shape, are associatedwith expanded capabilities for producing hydro-dynamic stabilizing forces (16). To understandhow fin erection alters its hydrodynamic per-formance, we constructed CAD models of thePacific bluefin tuna with erected and depressedsecond dorsal and anal fins and tested them witha computer fluid dynamics program for a rangeof speeds (2, 6, and 10 m/s) and yaw angles (1° to20°) (fig. S6). Erected fins had improved lift with-in a range of yaw angles from 1° to 8°, which

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Fig. 2. Biomechanics and hydrodynamics of tuna median fins. (A and B) Schematic of thehydraulic effect in the second dorsal fin (SDF) of tuna. (A) Depressed fin: compressible vascularchannels (CVC), fin rays (FR), expandable vascular channels (VC) between the FR, relaxed inclinatormuscles (IM), and incompressible chamber with vascular sinus (VS). (B) Erected fin: solid arrowsindicate collapsed CVC caused by contraction of the IM. Dashed arrow indicates outflow of liquid fromCVC to the VS and VC. (C) Isometric view of a CAD model of tuna shows the simulated flow patternassociated with the SDF and anal fin (AF) at 6 m/s and a yaw angle of 11°. (D and E) Top view of a tunaCAD model shows the velocity V, lift vector L, and drag vector D; a is the yaw angle formed by thefin plane and the velocity vector. (D) Enlarged view of the second dorsal fin without associated flow.

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increased with increased simulated speed (tableS5). This increment of lift resulted in improvedlift-to-drag (L/D) ratios, i.e., the best wing per-formance (18) up to 22 and 15% for the dorsal finand anal fin, respectively, at increased speedsfrom 2 to 6 m/s (fig. S6 and table S5). The L/Dincrement for both fins depressed at the sameconditions was 11%. This L/D ratio variation withwing sweep is comparable to the experimentaldata of morphing aquatic micro air vehicle (19).The improved performance of erected medianfins may be advantageous at turning maneuvers(20–22), which is in agreement with video analy-ses of swimming tuna (fig. S5 and tables S1 andS2). Both depressed and erected fins had maxi-mal L/D ratios within a small range of yaw angles,from 4° to 7°; this effect was reversed above 9°.As tuna are highly specialized in cruising, with arelatively large turning radius compared to otherspecies (23), this range may indicate possible op-timization of median fins for swimming controlat small inclinations from the axial movement.To determine the origin of the vascular system

within the fin sinuses, we performed vascularinjections, as well as histological, cytological, andimmunological assays. Injection of Microfil intothe VS revealed separation between the vesselsconnected to the VS and the arteries and veins(Fig. 3A and fig. S7). The Microfil was present onhematoxylin & eosin (H&E)–stained sections onlyin a subsection of vessels with veinlike morphology(Fig. 3A)—a characteristic of the lymphatic system(1, 7, 24). Pigmented aggregates resemblingmelano-macrophage centers (MMCs) were observed on theendothelium of the VS and connected vessels (Fig.3, A and B, and fig. S7). In other teleosts, MMCscontain lymphocytes and macrophages and havebeen shown to facilitate antigen presentation, sug-gesting a function similar to that of lymph nodesin other vertebrates (25, 26). Isolation of cells re-vealed that red blood cells (RBCs) were the dom-inant cells in the blood (>95%), whereas RBCsconstituted less than 40% of the cells in the sinusliquid (Fig. 3C); the rest of the cells had a lym-phocyte and myeloid morphology. The spleen infish is a lymphoid organ and considered to be thereservoir for the blood system (26, 27) and, as such,the leukocytes fraction in the spleen was higherthan in the blood but lower than in sinus liquid(Fig. 3C).To compare the different cell components of

the sinus liquid and blood, we used flow cytome-try (FACS, fluorescence-activated cell sorting) anal-ysis. FACS analysis revealed about 96% RBCs,1% lymphocytes, and 3% myeloid cells in blood,thus supporting our microscope observations. Bycontrast, the sinus liquid samples revealed a com-position of about 35% RBCs, 20% lymphocytes,and 40% myeloid cells (Fig. 3E). In addition,hematocrit was significantly lower in sinus liquidcompared to blood (Fig. 3D). Likewise, hemo-globin and mean cell hemoglobin concentrationswere reduced in sinus fluid, as was plasma lactatecontent (table S3). We used a LysoTracker markerto test whether leukocytes that were observed ona morphological level contained acidic vesicles.LysoTracker-positive leukocytes contain acidic

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Fig. 3. Vascular sinuses of median fins are connected to the lymphatic system. (A) Bloodand lymphatic vessels after blue Microfil injection into the VS of the anal fin. Internal lining of theperitoneal cavity (left), H&E histological section of the lining of the peritoneal cavity (right). Examplesof arteries, veins, and vessels staining positive for Microfil; melano-macrophage centers (MMCs) arelabeled. Bars, 1 cm (left panel); 150 mm (right panel). (B) Image of longitudinal section of VS. Veinsand MMCs in the endothelium of the VS are labeled; ruler in mm. (C) Cells were isolated from theVS, blood, and spleen. Upper panels, live cells; lower panels, methanol-fixed and Giemsa-stainedcells. Examples of the main three cell types are labeled: red blood cells, lymphocytes, and myeloidcells. Bars, 40 mm. (D) Hematocrit analysis of the VS liquid (left) and blood (right). Plasma, hematocrit(RBCs), and buffy coat (white blood cells and platelets) are labeled. (E) Cells were isolated fromblood (left) and the VS (right) and analyzed by flow cytometry using forward scatter (FSC) and sidescatter (SSC) for populations of RBCs, lymphocytes, and myeloid cells. (F and G) Cells were coculturedwith beads and analyzed by green fluorescence for phagocytosis and FL4 (unstained). Representativeanalysis of blood samples with gated positive cells (left) and VS (right). (G) Normalized analysisof fold change of cells positive for phagocytosis from three different experiments. *P <0.005,single-factor ANOVA.

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vesicles, which are typically associated with lyso-somal compartments in immune cells. LysoTrackerwas observed in myeloid cells at high levels andto a lesser extent in some of the lymphocytesbut was not observed in RBCs (fig. S7H). We thenperformed a fluorescent beads phagocytosis assayto test whether the leukocytes in the sinusliquid would function immunologically (24, 27).The percentage of phagocytic cells was higher(>4-fold) in the sinus fluid compared to the blood(Fig. 3, F and G), thus confirming the function-ality of the leukocytes in the sinus liquid. Tovalidate phagcyotosis, we performed a confocalmicroscopy three-dimensional analysis of beads

internalization, and engulfment and fusion withlysosomes of Staphylococcus aureus bioparticles(fig. S8). Cytoskeleton inhibition significantlydiminished phagocytosis of S. aureus, showingthat this process is an active function of phago-cytic cells in the VS fluid (fig. S8). Together, theresults suggest that the VS located at the base ofmedian fins is connected to the lymphatic sys-tem of tuna and filled with lymphatic fluid.We describe a musculo-vascular complex of

median fins in Pacific bluefin tuna, where theinclinator muscles and lymphatic vessels indi-cate hydraulic control of fins. We also found thiscomplex in other scombrids, including yellowfin

tuna, Pacific bonito, and Spanish mackerel, butit was absent in Pacific mackerel (Fig. 4A). Al-though the phylogeny of the Scombridae familytribes has been questioned (28), our finding cor-roborates a more recent whole–mitochondrial ge-nome analysis (29) showing the proximity of tuna,bonito, and Spanish mackerel tribes (Fig. 4B). Theapomorphy of the lymphatic network in medianfins may have evolved as a response to the demandfor swimming control in cruising specialist species,possibly related to pressure distribution aroundtheir bodies (30–32) (fig. S9). Differences in sec-ondary vascularization in scombrid fishes mayalso be related to variability in metabolic rates as-sociated with species specialization (6, 33). Con-stant fine adjustment of the sweep of the fins orwings can be observed in other animal classes andappears as part of swimming or flight control in-volved in lift-based propulsion (34). The discoveredmechanism described here may also inspire fur-ther development of biomimetic propulsive robotswith enhanced motion control.The use of fluids, in combination with muscu-

lar arrangements, for force transmission has beendescribed in a variety of invertebrate animals, in-cluding worms, mollusks, arthropods, and coelen-terates, as well as in penises in mammals (35, 36).Unlike other known examples of animal hydrau-lics, the musculo-vascular complex described herefor tuna fins is formed by the integration of thelymphatic vessels and muscles having skeletalsupport. The complex is composed of the threeelements of a canonical hydraulic system: musclesthat may serve as a hydraulic pump to pressurethe lymphatic fluid, vascular vessels to guide andcontrol the system, and fin rays acting as actuatorsto convert pressure energy into mechanical energy.

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Fig. 4. The lymphatic sinus in the Scombridae family. (A) Dissecting scope images of sectionstaken from the base of the second dorsal fin from Pacific mackerel, Spanish mackerel, bonito, andyellowfin tuna. The vascular sinus (VS) is labeled in Spanish mackerel, bonito, and yellowfin tunawith a black arrow but is not detected in Pacific mackerel. Bars, 1 mm. (B) Hypothesized developmentof the VS in the Scombridae family cladogram. The cladogram of the four scombridae tribes isreproduced with permission from a whole–mitochondrial genome analysis (22), based on 18 speciesof scombroid fishes. Hypothesized development of the vascular sinus based on the detection of VSin two species of Thunnini tribe (Thunnus albacares and Thunnus orientalis), one species of Sardinitribe (Sarda chiliensis), and one species of Scomberomorini tribe (Scomberomorus sierra) and thelack of the VS in one species of Scombrini tribe (Scomber australasicus).

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ACKNOWLEDGMENTS

We thank the Tuna Research and Conservation Center staff fortheir assistance with the tuna experiments, including C. Farwell andA. Norton. We also thank M. Ezcurra and the staff of the MontereyBay Aquarium for help in video recording of swimming tuna, andP. Chu, J. Tsai, I. Harel, and M. Dimitrov for technical assistance.High-speed video is courtesy of R. Kochevar, True Blue Films, andDiscovery Communications. Additional appreciation is extended toJ. Dabiri, J. Potvin, C. Lowe, D. Epel, C. Reeb, K. Palmeri, and N. Clarkefor useful discussions. Supporting data are in the supplementarymaterials. This work was supported by the Office of Naval Researchand Monterey Bay Aquarium. B.R. was supported by a postdoctoralfellowship from the Human Frontier Science Program Organization and

the NIH hematology training grant T32 HL120824-03. The authorsdeclare no competing financial interests.

SUPPLEMENTARY MATERIALS

www.sciencemag.org/content/357/6348/310/suppl/DC1Materials and MethodsFigs. S1 to S9Tables S1 to S5Movies S1 and S2Reference (37)

20 September 2016; accepted 15 May 201710.1126/science.aak9607

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Hydraulic control of tuna fins: A role for the lymphatic system in vertebrate locomotion

BlockVadim Pavlov, Benyamin Rosental, Nathaniel F. Hansen, Jody M. Beers, George Parish, Ian Rowbotham and Barbara A.

DOI: 10.1126/science.aak9607 (6348), 310-314.357Science 

, this issue p. 310; see also p. 251Sciencestiffen their dorsal fin. This provides extra stability during swimming.Triantafyllou). In bluefin tuna, a series of lymphatic vessels are integrated with muscles that allow the fish to raise andhomeostasis function has been co-opted to help facilitate dorsal fin rigidity and movement (see the Perspective by

show, however, that in the scromboid (tuna and mackerel) family of fish, this fluidet al.homeostasis. Pavlov immune response and−−The lymphatic system in fish has much the same function as it does in mammals

Hydraulic fins

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