The Blood Systems of Safoella and Spirographis. By D. W. Ewer (From the Zoology Department, University of Birmingham.) With 10 Text-figures. 1. INTRODUCTION. THE respiratory blood-pigment chlorocruorin is found only in sabellid, serpulid, and chlorhaemid polyehaete worms. The peculiarities and function of this pigment, together with the physiology of respiration and blood circulation in sabellids, have been studied by Fox and his collaborators (Fox, 1926, 1932, 1933, 1934, 1938; Eoche and Fox, 1933; E. F. Ewer and Fox, 1940). Before further work can be done on these lines it is desirable to have a more detailed knowledge than hitherto of the anatomy of the blood system in the animals concerned. It was for this reason that the present investigation was under- taken. A study has been made of the anatomy of the blood system of Sabella pavonina Savigny, and this has been compared with that of Spirographis spallanzanii Viviani. The latter species is so closely related to the former that Claparede (1868-71, p. 418) remarked: 'les jeunes Spiro- graphes sont done de vraies Sabelles.' 2. PREVIOUS WORK. The anatomy of the blood system of Sabella was studied by Milne Edwards (1838). He described the arrangement of blood-vessels which could be seen by means of a dorsal dissec- tion. In the same year Grube (1838) gave an account of the anatomy of the blood system of Spirographis. His descrip- tion was based on the observation of living material and the dissection of fixed specimens. He made accurate observations on the movement of the blood in the crown. Both authors 588 D. W. EWER described a dorsal vessel running over the surface of the gut, though Grube was very doubtful about the length of this vessel. They both described the ventral vessel with the curious lateral coilings of the paired ring vessels which lead out of it in each segment. Grube described the vessels of the thorax, and, with particular accuracy, the two longitudinal lateral vessels in the abdomen. In 1873 Clarapede redescribed the anatomy of the blood system of Spirographis. He appears to have used only fine hand-cut sections. He was able to show that the earlier authors had been wrong in describing a dorsal vessel over the surface of the gut. He showed that the gut is surrounded by a sinus, replaced anteriorly by a plexus over the oesophagus. He stated that not only the vessels to the crown but also the ventral vessel arise from this plexus. Jaquet (1886) re-examined the blood system of Spiro- graphis. He rediscovered the paired lateral vessels which had been missed by Claparede. In his description of the thorax he followed that of Claparede in every detail. Meyer (1888) once again described the blood system of Spirographis. He figured and described only the main vessels. He confirmed' the existence of a gut sinus, and showed that over the oesophagus the sinus concentrated to form a dorsal vessel which carried the greater part of the blood from the gut sinus to the crown. The blood from the crown returned by definite circum-oesophageal vessels to the ventral vessel. Meyer does not draw or describe the plexus found by Claparede around the oesophagus, but it is apparent from his general description that he was aware that it was there. De Saint-Joseph (1894) published a description of Sab el la in which he confirmed the presence of a gut sinus and also of lateral vessels. He did not describe the thorax. There have been a number of descriptions of the structure of the crown filaments, which have been reviewed by Nicol (1930), whose paper includes an accurate description of the structure of the vessels of the filaments and pinnules of Sabella. The accounts of the blood systems of the animals described as Sabella alveolata by Williams (1851) and of Sabella BLOOD SYSTEM OF SABEUA 589 arenilega by Cosmovici (1879) have sometimes been com- pared with descriptions of Sabella pavonina. They refer, however, to Sabellaria alveolata (L.) and Bran- chiomma vesiculosum (Montagu), respectively. 3. MATERIAL AND METHODS. Sabella was sent to Birmingham from Plymouth, and from Millport in Scotland, while Spirographis was collected at Eoscoff in Brittany.1 Paraffin sections cut to a thickness of 10 and 20ft were used. Material was fixed in Bouin-Hollande, Duboscq-Brazil, Schau- dinn, Champy, Flemming, Hermann, and Camoy. The fixative described by Keilin (1920) was also used. Of these the best results were obtained with Duboscq-Brazil for Sabella and with Keilin's fixative for Spirographis. Sections were stained in Heidenhain's iron haematoxylin, Delafield's haematoxylin and orange G or eosin, Mallory's triple stain, and Mann's methyl blue and eosin. A modification of the method of staining blood by the benzi- dine reaction described by Pickworth (1934) was used. Whole worms were fixed in a 4 per cent, solution of formaldehyde in sea-water for two days and then washed repeatedly in sea-water to remove the formaldehyde. The material was afterwards stored in sea-water saturated with thymol. Material thus preserved has retained its staining power for at least a year. When required the material was washed in running water and then sectioned on a freezing microtome and stained as described by Pickworth. Thick sections of 100-250/u. were used. Small worms were stained by the modification of the benzidine technique described by Slonimsky (1927). Living worms were first washed in tap water before being treated with the reagents, as sea-water produces a yellow incrustation. This method has the disadvantage of being useful only for small and living specimens. The reagents will not penetrate into large specimens. Small living worms were used to observe the directions of flow of the blood through the transparent tissues. 1 My thanks are due to the Director and StaS of the Laboratoire Lacaze- Duthiers at Roscoff for their hospitality and help. NO. 328 Q q 590 D. W. EWER 4. THE ANATOMY OF THE BLOOD SYSTEM OF SABELLA. The blood system of Sabella will be described in detail and those differences which are found in Spirographis will then be given. The body of Sabella is divided into two regions; anteriorly the head and thorax, composed of a variable number of seg- ments from six to twelve, and posteriorly the abdomen, with segments up to the number of about three hundred. In describing the anatomy of the blood system it is convenient to start with the abdomen. 1. The Abdomen. The general organization of the blood system in the abdomen may be seen by an examination of Text-figs. 1 and 2. The vascular system in the abdomen is organized into (a) the longitudinal vessels—gut sinus, paired lateral vessels and ventral vessel—and (b) the circular vessels of each segment—ring vessels, lateral connective vessels, segmental dorsal vessels, notopodial vessels, neuropodial vessels, trans-septal vessels, and ventral gland-shield vessels. The Gut Sinus (or peri-intestinal sinus).—In a freshly dissected specimen or in a small transparent worm the gut sinus is immediately recognizable (Text-fig. 1, g.s.). The blood can be seen to be carried forward by the peristalsis of the sinus wall. The sinus runs the whole length of the worm from the hindmost segment to the anterior septum of segment III. In each seg- ment it receives a pair of ring vessels (Text-fig. 1, r.v.) which come into it ventro-laterally. The Ventral Vessel.—The ventral vessel (Text-fig. 1, v.v.) runs backwards along the whole length of the worm from the posterior portion of segment II. It lies in the ventral mesentery above the giant fibres and nerve-cord (Text-fig. 3, v.v.). The vessel is circular in cross-section and may reach a diameter of 100/L*. It is covered with black cells (Text-fig. 3, &.C.).1 From the ventral vessel arise a pair of ring vessels as 1 These cells containing black granules have been called 'chloragogen as.. s.dv d.mb L.v U so D ! GO H H neuc) tieu v. CD vm.b. W H rv. v.v. TEXT-FIG. 1. Sabella pavonina. Diagram of the vessels in the abdominal segments seen from the posterior aspect. The body- wall and septa of the right side, and the mesenteries, have been out away. Capillaries are omitted, d.m.b., dorsal muscle block; g.s., gut sinus; I.e., lateral connective vessel; l.v., lateral vessel; neu.g., neuropodial gland; neu.v., neuropodial vessel; not.g., notopodial gland; not.v., notopodial vessel; r.v., ring vessel; a.d.v., segmental dorsal vessel; t.s., trans-septal vessel; v.g.s., ventral gland shield; v.g.s.v., ventral gland-shield vessel; v.m.b., ventral muscle block; v.v., ventral vessel. 592 D. W. EWER lateral branches immediately in front of each septum (Text- fig, l.r.i?.; Text-fig. 3, r.v.). The Lateral Vessels.—These vessels (Text-fig. 1, l.v.) can be seen through the skin in small living worms as two zigzag lines running along the whole length of the body in a latero-dorsal position. They run forward as far as the posterior septum of segment II. The lateral vessels are joined to the ring vessels in each segment by two lateral connective vessels (Text- fig. 1, I.e.). On each side of each segment the lateral vessels give off two blind-ending branches—the segmental dorsal and the notopodial vessels (Text-fig. 1, s.d.v. and not.v.). If one of the lateral vessels is followed forwards along the dorsal muscle block beneath which it runs, its relations may be understood.