19. Ontogeny of thymus in red sea bream, Pagrus major

VI

staining, phagocytic activity, autonomous fluorescence and other observations described below,we classified hemoeytes of the ascidlanHalocynthia roretzi into ten types; large amoeboeyte (30-40%: vacuolated cell, Fuke 1979), vesicular cell (15-27%), granular cell (2-7%), minute granular cell (12-23%), macrophage (7-12%: fine granular amoeboeyte, Fuke 1979), morula cell (5-10%), large hasophilic cell (1-2%), large acidophilie cell (< 1%), single granular cell (13%), lymphocyte (1-5%). In a 5ram EGTAsolution (in 1/2 M NaCI, pH 5.8-6.0), morphological differences among the types were most recognizable under phase contrast microscopy, and only large amoebocytes (LA) and macrophages spread on glass. LAw'as small and had an irregular shape before spreading. LAwas considered as the encapsulating cell, because LAspread rapidly and widely on glass and several LAs fused into a syncytium in the presence of Ca ++. We also found that NH4CI, causing morphological change in several types of cells, was a useful tool in cell identification. Previously reported monocional antibody LIB15 stained only minute granular cells. 17. COLONY SPECIFICITY IN THE COMPOUND ASCIDIAN, APLIDIUM YAMAZII. Yasuho Taneda and Yuki Kawana. Faculty of

Education, Yokohama National University, Ho. dogaya-ku, Yokohama 240, Japan. In some species of compound ascidians, a phenomenon called colony specificity is well known. This phenomenon has been studied mainly in botryllid ascidians with a common vascular network in the colony. Although there is no common vascular system in the colony,Ap"/u//umyamaz//shows apparent colony specificity. We revealed the following characteristics of colony specificity in A. yamaz/i. Contact between two fragments of the same colony always led to fusion, even if they made contact with growing edges or cut surfaces. Contact with growing edges between two incompatible colonies led to nonfusion reaction (NFR). NFR was characterized by the concentration of test cells at the contact area and fragmentation of the tunic. Four types of test cells could be recognized in the tunic. Granular cells with filopodia seem to be the main components of test cells. In the case of contact with cut surfaces between two incompatible colonies, they fused in spite of incompatibility. Several days after contact, they separated without concentration of test cells. Therefore, the cuticle of the tunic plays an impor-

Abstracts

tant role in NFR. 18. THE PRIMITIVE COMPLEMENT SYSTEM IN AGNATHA: CHARACTERIZATION OF ZYMOSAN-BINDING PROTEINS OF HAGFISH SERUML Tamotsu Fujil, Aya Sekizawa* and Susumu Tomonaga**. H/ro-

shima Women's University, Hiroshima 734,

*Seitoku University, Matsudo 271 and **School of AUied Health Sciences, Yamaguchi University, Ube 755, Japan, The zymosan-binding proteins of hagfish serum (HZBP) were analyzed by immunoeleetrophoresis (IEP) and SDS-PAGE. Zymosan particles were treated with normal serum of the hagfish, Eptatretus burgeri, for 1 hr at 10*C. The serum-treated zymosan was washed with PBS followed by incubation with 1% SDS in PBS(pH7.5). The HZBP were effectively eluted from washed zymosan by subsequent treatment with an alkaline buffer(pH 11.0) containing 0.1M hydrazine. Upon IEP, the greater portion of eluted proteins formed a single precipitin line in the slow fl-globulin region against rabbit antiwhole hagfish serum as well as anti-HZBP. SDSPAGE analysis of eluted proteins revealed an intense migrating major band (HX) with a MW of 170 kDa. After reduction, the HX migrated as two proteins with MWs of 98 kDa and 77 kDa. The I-IX, however, was not eluted from zymosan incubate.d with serum which had been pretreated with methylamine, EDTA, or heated at 42"C for 30 min. These results suggest that there is an alternative-like pathway of complement, and that the HX represents the C3 of the hagfish. 19. ONTOGENY OF THYMUS IN RED SEA

BREAM, PAGRUS MAJOR. Rlichi Kusuda, Masaru Ikemoto and Hideaki Enzan*. Fish

Disease Laboratory, Faculty oyAgriculture, Kochi University, Nankoku, Kochi 783, Japan, *Depart. ment of Pathology, Kochi Medical Schoo~ Nankok~ Kochi 783,Japan. Theontogeayofthe thymus in Pagrus major from day 0 to 2 months after hatching was studied morphologically. The thymic anlage first appeared within the pharyngeal epithelial cells above the third gill arch at the 12th day after hatching. The parencbyma consisted of small and large lymphoeytes, having a diameter of about 4 and 6 #m, respectively. The thymus at this stage was separated from the surrounding mesenchymal tissue by epithelial reticulum cells and a basement membrane. Later, small lymphoeytes became dominant in the outer peripheral portion while large lympho-

Abstracts

cytes were in the inner portion. Macrophages were observed among the lymphocytes. By 40 days after hatching, thymus differentiated into outer and inner zones: cortex and medulla. At this time blood vessels invaded from the surrounding connective tissue into the thymus. Lymphocytes were also observed in the connective tissue around thymus. From these results the basic structure of the thymus was formed by 40 days after hatching.

VII

phocyte marker in humans and most mammals. We investigated the characterization of carp peripheral leucocytes that form spontaneous rosette with red blood cells of sheep (SRBC), horse (HRBC), bovine (ORBC), and rabbit (RRBC). The frequency of E-rosette formation with SRBC and HRBC was higher than that of ORBC and RRBC. E-rosette formation of carp leucocytes depended on the incubation time and occurred at a temperature ranging from 0"C to 37°C. Variations in surface morphology of re20. ANTIGEN RECEPTOR ON LYMPHO- sere forming cells were observed under the scanCYTES AND MACROPHAGES IN YEL- ning electron microscope as for mammalian lymLOWTAIL, SERIOI~ QUINQU~tADIAT~ phocytes. Rosette forming ability of nylon fiber Masaml Hamaguchl and Rllchi Kusuda*. adhered cells was stronger than that of nylon fiber Nansei National Fisheries Research Instiaae, non-adhered cells. The presence of surface imOno-cho, Hiroshima 739-04, Japan and *Fish munoglobulin among carp peripheral leucocytes DiseaseLaboratory,FacultyofAgriculture,Kachi that forms E-rosettes was proved by the imUniversity,Nankoku, Kochi 783,Japan. Effectof munofluorescence technique, although rosette various antibodies on erythrocyte rosette forma- formation was not dependent on surface Ig. tion by peripheral lymphocytes and peritoneal These results suggest that spontaneous E-rosette maorolShages in unsensitized yeliowtail was inves- formation could not be applicable as a routine tigated. Rabbit red cell (RRBC) and sheep red technique for the classification of carp leublood cell (SRBC) were used. RRBCs were pre- cocytes, unlike mammalian lymphocytes. treated with yeUowtail serum and SRBCs were pre-treated with neuraminidase. Monoclonai 22. CHARACTERISTICS OF CELLS INFILantibodies (Mab) prepared and examined for the TRATING INTO CHV-INDUCED CARP rosette formation-inhibition test were anti-thy- PAPILLOMA. Natsumi Morita and Tokuo mocyte cell surface antigen, anti-macrophage cell Sane. Laboratory of Aquatic Pathology, Tokyo surface antigen, anti-yellowtail Ig and anti-yel- University of Fisheri~, Minato-ku, Tokyo 108, Iowtail complement C3. Anti-lymphocyte and Japan. Carp herpes,Arus (CHV) induces papilanti-macrophage polyclonal antibodies were also loma on carp (Cypfinus carp/o L.), and PBL may made and examined. RRBC rosette formation play an important role in regression. The regreswas inhibited more than 90% by treatment with sion period of CHV-induced tumors appeared in anti-complement C3 Mab, but less than 20% by carp injected with anti-carp PBL serum. In this anti-Ig Mab, suggesting that receptors for com- study, we attempted to characterize cells that plement C3 and Ig are important factors for infiltrate into tumors. Upon uitrastructural obbinding RRBCwithyellowtailcells. On the other servation, two types of granulocytes were recoghand, SRBC rosette formation was inhibited by nized in the papillomatous tissues. One type of pretreatment of lymphocytes and macrophages granulocyte possesses numerous spherical and with anti-Ig Mab. Anti-lymphocyte and anti- high electron-danse granules, and in the other, macrophage Mabs showed no significant inhib- the granules vary in shape with irregular density. itory effect. This indicates that antigen-receptor, Cells harvested from the papillomatous tissue which may be Ig receptor itself or Ig receptor-like were fractionated by percoll gradient density cenmolecule, plays a major role in the binding be- trifugation. In the density of 1.075-1.121g/ tween SRBC and yellowtail cells. ml(F.3) and higher than 1.121g/mi as F.4, granulocyte-like cells were predominant. Granulo21. CHARACTERIZATION OF ROSETTE cyte-like cells in both F3 and F.4 revealed a FORMING CELLS IN CARP PERIPHRAL round shape with eccentric nuclei and were negaLEUCOCYTES. Tadatoshl Kitao and tive for either peroxidase or alkaline Terutoyo Yoshida. Department of Fisheries, phosphatase, but positive for acid phosphatase. Faculty of Agriculture, Miyaz_~ University, Eosinophilic granules were also found in cells of Miyazaki889-21,Jatmtt Erythrocyte (E)-roset te F.3, but granules in cells of F.4 were equivocal. formation has been widely accepted as a T lyre- However, these cells were considered to be sirni-