eaethy Tait preverere tees tay shea aate ff i tree rie Cerariii A egeaecesseasbent cee Rede nae Boe! ee ee Gar rears inert eltan pea ted ft ayy, a De ryes a het Bawa b ae Se mrhssed loses NOH ly eft ieh yy ¢ peat Peed oetherte: ae ars : of ; is yagi . ory : ise srebpiece ne vi PAS oderenee ma : : eat! ROIS TTA TES ba t r : ls ssi Ae pena nage bigaet a SEsee Peashstatodeter stabs se int Digitized by the Internet Archive in 2009 with funding from Ontario Council of University Libraries http://www.archive.org/details/anatomicalrecord12bard THE ANATOMICAL RECORD EDITORIAL BOARD Invinc HarpEstTy WaRREN H. Lewis Tulane University Johns Hopkins University CLARENCE M. JACKSON Cuar.ies F. W. McCiure University of Minnesota Princeton University Tuomas G. LEE WiuraM 8S. MILLerR University of Minnesota University of Wisconsin Freprric T. Lewis FLORENCE R. SABIN Harvard University Johns Hopkins University GrorGE L. STREETER University of Michigan G. Cart Huser, Managing Editor 1330 Hill Street, Ann Arbor, Michigan VOLUME 12 FEBRUARY-MAY RY-MAY, 1917 so A CS \% a» : oe ae: PHILADELPHIA THE WISTAR INSTITUTE OF ANATOMY AND BIOLOGY ‘Nath oye ia 2 * clas gi of @L ‘ aig wal a nm A a SO; © uo me i AS +s : 2 ee be. a: eh ne me Ae CONTENTS No.1 FEBRUARY Frank E. BuaispELu, Sr. The anatomy of the sacro-uterine ligaments. Twenty-two BANEES.. . 2.2 ete cael Se oer ee aeege iz Mage ete = «= 2 os AR TAC oe iL ARTHUR WILLIAM Meyer. Spolia anatomica, addenda II. Twenty-two figures....... C. M. VanperBurcH. The hypophysis of the guinea pig. Seven figures............... H. A. Cuarrensoure. The thoracic duct of the adult guinea pig. Thirty-four figures. . ELBERT CLARK AND RuskIN H. LHamMon. Observations on the sweat glands of tropi- SERS INGER eT enaeeN. PW MPUITCS, . 6. olot ceca as os eee s ne odes cbc oesne ncacsaun Gotper L. McWuorrer. The relations of the superficial and deep lobes of the parotid gland to the ducts and to the facial nerve. Two figures..................2.0..000- Sara B. Conrow. Further observations on taillessness in the rat. Three figures..... Montrose T. Burrows. The significance of the lunula of the nail. Two figures...... B: TatBot Stroneman. A preliminary experimental study on the relation between mitochondria and discharge of nervous activity................ cece cece cccccccee T. Srantey O’Matiey. An anomalous Vena pulmonalis within the lung. Two figures EBEN Carey. The anatomy of a double pig, Syncephalus thoracopagus, with especial consideration of the genetic significance of the circulatory apparatus. Ten figures. Rosert T. Hance. A device to increase the efficiency and ease of manipulation of the ime adjustment of the microscope. One figure...............0ccc ccc cece ccc cece No.2 MARCH* Orto C. GraserR. On the mechanism of morphological differentiation in the nervous system. Il. The relation between compression and the development of a series of wemeles. Tight text figures and three plates... .........060 00s .icecceecccccscesccase JoHN SunpwatL. The choroid plexus with special reference to interstitial granular Se IRR PEST SVIR EE On RM eh os rhs Gis Iui aie ak pk PRR aM RSS x Ae RRR «disses Yc alia. clive oiloclonwe F. E. Cuipestrer. Intestinal hernia in two species of frogs. Three figures............. J. M. Srorsensurc. Observations on the influence of isolated ovaries on the body growth of the albino rat (Mus norvegicus albinus). Two charts................... FRANKLIN P. REAGAN, Epwarp E. MacMortanp, and Sruarrt Mupp. Anterior haem- atopoesis in chemically treated teleost embryos under continual observation. Eleven RARE ED Mees eM a, YO Sree ae Sas Uae Rtg ws « Oui dks aikins Dace Rae ta de GEORGE W. Corner and A. E. AMsBauaH. Oestrus and ovulation in swine............ ARTHUR WILLIAM Meyer. Intra-uterine absorption of ova. Seven figures .. J. B. Jounston and Epna G. Dyar. Methods of mounting sections in gelatin ......... Louis H. Kornver. The value of absolute alcohol for removing adherent paraffin sec- Prsens paper OF pastebogrd tray... ~:~ < «dens. -~-v a oUakw is Seedy hoveu tea vs Epmonp Soucuon. Preservation of anatomic dissections with permanent color of mus- cles, vessels and organs. A supplementary note, describing another method, the MRI isk... 2. shes hy MENS a vel ers Poca em ne biotin as Re oc es en aes W. D. Wat.is. The development of the human chin:......................eecceeeeeee Ricuarp W. Harvey. Notes on two cases of anomalous right subclavian artery ....... ill ei 43 95 113 “= iv CONTENTS No.3 APRIL Georce K. Hasurpa. The lymphatics system of the guinea-pig. Nineteen figures..... 331 Apo.tr H. Scuvyrz. Ein Paariger Knocken am Unterrand der Squama occipitalis. Mit 2 figuren im text........ 2-2-2. eee e eee erect eee ene eee eee eee e ses sensscees 357 G. E. McCiune. Cooperative technique........-...-----+-0++-++00-+- += ensue eens 363 Sana B. Conrow. A six-legged rat. Five figures.....-...-.:«-.-.-.-+---->s=6eee 365 Ropert T. Hance. The fixation of mammalian chromosomes. Two text figures and three plates... ......--- 02-2. 2e cece cece eee een cee eases sencs te cng ee sns n= see 371 F. E. Cawwester. Hermaphroditism in Fundulus heteroclitus. Three figures.......... 389 C. V. Morritu. Reference model of the thoracic viscera. Three figures............... 397 C. CG. Macxirx. Notes on the preparation of bones from madder-fed animals.......... 403 Ciarence L. Turner. A culture medium for Euglena with notes on the behavior of ch CY i en MEE 8 407 No.4 MAY G. B. Wistocxi. The action of vital dyes in teleosts................-........--se eee 415 Hat Downey. Reactions of blood and tissue cells to acid colloidal dyes under experi- mental condition. Four figures (one plate). ..... 1. 6.0.06 .e ence ess- + >=: =p 429 James H. Warren and JonatHAN ForMAN. Observations on the occurrence of eosifo- philic leucocytes and the granule cells of Paneth in the vermiform appendix of man. 455 Lewis H. Weep, An anatomical consideratian of the cerebro-spinal fluid.............. 461 From Mi. . Variations . Experiments THE ANATOMY OF THE SACRO-UTERINE LIGAMENTS FRANK E. BLAISDELL, SR. the Laboratory of Surgical Pathology of the Medical School of Stanford . Introductory.... . py Miethods«ot study andmaterial .....c0c ex oe oh oe . Brief review of the literature........:........-..... BOUT ONOLOPICHlmes eae eee Macon s. ef Pac clece dees University TWENTY-TWO FIGURES CONTENTS b. Descriptive and topographical.................... 1. Guinea pig Cr em &W bo Primate series. . Pxplamatvory remarks...s2.1..20 0at-. a. ss se> . Comparative series.......... (Cavia cabayia).......... . Belgian hare (Lepus europaeus)................. Cat (Helis dontesticus). ar 2 <5-1<«%!<', -- PRON GATS TAININ ARIES «fae ichce wis sc otek bP CrenenslcCONSIC CTALIOUS: 24 .4< 2s. vracia + «'> os ixaiare iaVonkceys (MaAcHeus))-. 2 ane wccs as wees oks Pees Ue iiehibool: vetaueye a. Examination of the structures in a recent post-mortem b. Histological observations. ‘Fixed material....... _ . Adult 35 years old 2. Child 10 years old 3. Infant 40 days old 4. Adult 65 years old 5. Adult 83 years old THE ANATOMICAL RECORD, VOL. 12, No. 1 FEBRUARY 1917 . The musculi levatores uteri . Discussion . Summary and conclusions . Literature cited 2 FRANK E. BLAISDELL, SR. I. INTRODUCTORY A great deal has been written upon the anatomy of the pelvic floor and the mechanical supports of the pelvic viscera. While all of the contributors have more or less advanced our knowledge of the anatomy of the pelvis, all have failed to completely elu- cidate the anatomical and physiological importance of the fibro-elastie tissue that forms a more or less dense mesh-work, filling up the interval between the peritoneum above; the pelvic diaphragm covered by its facial sheath below; the obturator fascia and periosteum of the innominate bone laterally; the bladder, vagina, and uterus with their fibrous investment medi- ally; and finally, the fascia over the sacral promontory, sheath of the sacral plexus, and fibrous tunic of the rectum, posteriorly. Cameron and Moritz have made the nearest approach to the proper solution of the question, but they failed in not subjecting the ‘compact mass’ filling up the interval between the perito- neum and pelvie floor as above limited to the proper anatomical analysis. The primary object of the present investigation is the study of the so-called sacro-uterine ligaments; to determine what they really are and to establish their relation to the transverse liga- ments of the uterine neck, recto-uterine muscles and recto- uterine folds. Il. METHODS OF STUDY AND MATERIAL From a survey of the work which has been done, it is evident that a comparative and a more comprehensive study is abso- lutely necessary for the proper solution of the problem. In ac- cordance with that view, cadavers that had been embalmed in the usual manner for dissection, and others preserved with 10 to 40 per cent solutions of formaldehyde, and fresh material in and from the autopsy room, were used. Recent or unfixed material has the advantage of being perfectly flexible, permitting of ex- perimental procedures not possible on the firm and _ inelastic material of the dissecting room. Finally, a careful study of frozen sections revealed some most interesting points. THE SACRO-UTERINE LIGAMENTS 3 The human material included fetuses, the new born, and cada- vers of the following ages: 40 days, 10, 24, 35, 65, and 83 years. The animals most readily available were the guinea pig (Cavia cabayia), Belgian hare (Lepus europaeus), cat (Felix domes- tica), dog (Canis familiaris), and monkey (Macacus). In the whole series the parametrium containing the so-called ligaments and folds were cut in serial sections and subjected to a careful examination. Experimental observations on the pelvic organs were made on the animals while under an anesthetic. Since the appearance of a preliminary report of this paper in June, 1913, there has been published an interesting paper by Moritz (714), in which the author considers the distribution and significance of the parametrium. His method has been to obtain the pelvic contents as soon after death as possible, by having them removed right down to the bone and the pelvic floor cut away with them. The specimens were then fixed in 10 per cent solution of commercial formalin. Not ‘dissected,’ in the ordinary acceptation of the term, but sections were cut in varying planes and directions. Ill. BRIEF REVIEW OF THE LITERATURE a. Chronological The more important contributors are the following: Kocks, (1) (’80), Derry, (10) (’07), Hart, (2) (’80), Paterson, (11) (’07), Mackenrodt, (3) (95), Ovenden, (12) (’07), Holl, (4) (97), Cameron, (13) (07-08), Harman, (5) (’98), Smith, (14) (’08), Thompson, (6) (00-06), Fothergill, (15) (’08), Deaver, (7) (03), Somers and Blaisdell, (16) (713), Montgomery, (8) (’03), Moritz, (17) (’14). Stony, (9) (04), Kocks (1) described the cardinal ligaments. Mackenrodt (3) de- fined the ligamentum transversale colli. Deaver (7) states that the false posterior or recto-uterine ligaments are composed of two peri- toneal layers which pass backward from the posterior surface of the uterus and vagina to the upper portion of the rectum, forming the lateral boundaries of the pouch of Douglas; external to each are found the true or muscular utero-sacral ligaments, which are flat muscular bands that extend from the uterus at the level of the internal os, to the sides of the sacrum, passing beneath the layers of the recto-uterine ligaments. 4 FRANK E. BLAISDELL, SR. Montgomery (8) says that the utero-sacral ligaments, while consist- ing of folds of peritoneum, also contain muscle fibers, which are de- rived from the superior muscular layer of the uterus. Paterson (11) considered the suspensory ligaments. Ovenden (12) reviews Mackenrodt’s work and states that the ligamentum transver- sale colli is worthy of being recognized as a distinct ligament, and that the utero-sacral ligament blends with the former near its insertion into the uterus. Cameron (13) considers the utero-sacral igaments as a part of the general perivascular mass. Moritz (17) in criticizing Dr. Ovenden’s views states: “I hope to demonstrate that the sketch she published of the insertion of the liga- ment (transverse of the neck) shows clearly that the so-called hgament is simply a portion of the parametrium containing the uterine artery, artificially separated from the surrounding mass.” He agrees that the parametrium constituting the ligamentum transversum colli is of great importance physiologically, in that they fix the cervix and form the most fixed point of the uterus. In a brief summary he states that: “the whole structure is nothing but an inseparable continuation of parametrium which surrounds and fixes the cervix.” He considers the utero-sacral ligaments as small folds of peritoneum, and contain nerves, perineural connective tissue and smooth muscle; they carry the nervi erigentes, pushing their way through the mesodermal tissue from their points of emergence from the anterior sacral foramen. b. Descriptive and topographical The descriptive and topographical statements by Cunningham (19) of the connections of the uterus and its relations to the peritoneum laterally and posteriorly may be taken as the consensus of opinion among anatomists. The deep pouch between the uterus and vagina in front and the rectum behind is called the pouch of Douglas (exca- vatio recto-uterina) and its entrance is bounded on each side by a erescentic peritoneal fold, which passes from the posterior surface of the cervix uteri to the posterior wall of the pelvis, and ends near the side of the rectum. These crescentic folds are called the recto-uterine folds (plicae recto-uterinae), and each contains between its layers a considerable amount of fibrous and smooth muscular tissue. Some of these fibers, which are continuous with the uterine wall, pass backward to reach the rectum and constitute the recto-uterine muscle (liga- mentum sacro-uterinum). In many cases the recto-uterine folds be- come continuous with one another across the middle line behind the cervix uteri, and form a transverse ridge termed the torus uterinus. A comparison of Morris (20), Piersol (21), and Gray (22) with Cunningham (19) and others, inevitably leaves the student in doubt as to what really constitutes a sacro-uterine ligament and its rela- tion - only to the plica recto-uterina but the musculus recto-uterinus AS Well. THE SACRO-UTERINE LIGAMENTS 5 Morris (20) calls the posterior peritoneal folds of the uterus, ‘the recto-uterine ligaments,’ and states that they become continuous with the peritoneal investment of the second part of the rectum, and that between their layers lie the utero-sacral ligaments, the latter being flat fibro-muscular bands, extending from the highest part of the cer- vix uteri, where they are more or less continuous with the uterine fibers in the recto-uterine peritoneal folds, to the sides of the sacrum f fi 7 ; ‘Bey ff as. Y at ‘ - wi use te Ureter__ 3 ; i y Fs P vy * Ureter : KK A s Pt ‘| mY i ; 7 |. — Ovarian Artery & 7 a .* Veins ‘ : 7 ; , 7 Pi ay / ; . i! f Hypogastric Arle ry semgay ralor yy Arte ry ft ' ——, S » ie Plica Sacro-Uterina- Va inal Vea Mee : Torus Uterina__. Fig. 1 Semi-diagrammatic drawing of female pelvic viscera viewed from above, uterus and adnexa being drawn forward and vessels projected against peritoneum. opposite the lower border of the sacro-iliac articulation. They run one on each side of the rectum near the junction of the first and sec- ond parts. Their muscle fibers (the recto-uterine muscles) become continuous with those of the rectum posteriorly and more anteriorly they lie in the recto-uterine peritoneal folds, which form the lateral boundaries of the pouch of Douglas. Piersol (21) states that between the layers of the folds (plicae- recto-uterinae) robust bundles of fibrous and smooth muscular tissue 6 FRANK E. BLAISDELL, SR. extend from the uterus to be inserted partly into the rectum, these constituting the utero-rectal muscle, and partly into the front of the sacrum as the utero-sacral ligament. Gray (22) says that the sacro-uterine ligaments (plicae-recto- uterinae) are contained in the peritoneal folds of Douglas. They pass from the second and third bones of the sacrum, downward and forward on the lateral aspects of the rectum to be attached one on each side of the uterus at the junction of the supra-vaginal cervix and the body. They contain fibrous tissue and unstriated muscle fibers. Muscular fibers from the uterine wall to the rectal wall constitute the recto-uterine muscle. This muscle is part of the sacro-uterine igaments. IV. EXPLANATORY REMARKS The terms cardinal ligament, ligamentum transversale colli uterini and suspensory ligament are synonymous. The para- metrium includes all of the fibro-elastic tissue lying lateral to the uterus; the paravaginal tissue that which lies opposite the vaginal vault; while the term paraplical will likewise signify the tissue lateral to the recto-uterine folds, or more specifically that lying lateral to the recto-uterine fossa and fibrous tunic of the rectum. It fills the subperitoneal cavity as defined by Moritz. The sacro-uterine and recto-uterine folds are the same struc- turally, the only difference being the manner of termination, which makes it necessary from a physiological standpoint to speak of them separately. The term fibro-elastie will be used instead of Cameron’s (13) term perivascular tissue, because the term perivascular defines only a part of the parametrial supporting tissue, and its func- tion is not primarily to support blood vessels as will be ex- plained later. V. COMPARATIVE SERIES In animals, which habitually assume the horizontal position in locomotion, there is to be found a simpler and less condensed condition of the uterus as regards bulk than in those animals which assume the erect position. Such animals possess either two uteri or a uterus bicornis. The uteri are therefore com- paratively less bulky and the weight is distributed along a more extensive peritoneal attachment. Such attachment usually ex- THE SACRO-UTERINE LIGAMENTS 7 tends as far cephalad as the caudal pole of the kidneys. At all times, except during short periods of time when other than the horizontal position is assumed, the weight of the uterus, dis- tended bladder and rectum, cause these organs to fall ventrad in the hypogastric region of the abdominal cavity. The uterus in a state of physiological rest is relatively light and conse- quently there is less need of true or fibrous ligaments, such as are present in the higher Primates, where the uterus is more con- Kidney Rectum ___ Uterus _ Bladder Fig. 2 Photograph showing relations of rectum, bladder and extensive peri- toneal attachment of uterus in an animal which assumes the horizontal position in locomotion. Dog (Canis familiaris). centrated in bulk and weight, and the erect position is assumed habitually, except during limited periods. Therefore, in animals possessing a uterus bicornis, the peri- toneal reflections are sufficient with a very meager amount of parametrial tissue to maintain the normal position of the or- gan. In the small series of animals examined during the prepa- ration of this paper, one very important and significant fact was determined, and that is, that there were always bundles of smooth muscle fibers present in the peritoneal folds passing dorsad from the vagina to the rectum. Laterally, in the para- metrial and paravaginal tissue, such bundles were practically 8 FRANK E. BLAISDELL, SR. absent and the fibro-elastic bundles were reduced to a mini- mum. It is therefore logical from an evolutionary standpoint to conclude that the presence of the recto-uterine muscle in these folds appears at an early date in the vertebrate series, and that they possess a phylogenetic significance. 1. GUINEA PIG In the guinea pig, two distinct peritoneal folds pass dorsally from the sides of the vagina to the sides of the rectum. They Excavatio pararectalis _ es ia ( Plica recto-vaginalis Plicavascularis _/ J : rae | Excavatio recto-vaginalis le Ligamentum latum \4A Fig. 3 Drawing showing rectum, vagina, uterus, recto-vaginal pouch and recto-vaginal folds in guinea pig (Cavia cabayia). are therefore recto-vaginal folds, and must be termed the lat- eral folds to distinguish them from the median recto-vaginal reflection of peritoneum at the bottom of the recto-vaginal fossa. Lateral to the rectum and recto-vaginal fold is the para- rectal fossa, which is in turn bounded laterally by a peritoneal fold conveying utero-vaginal blood vessels. The lateral recto- vaginal folds and broad ligaments of the uterus are extremely thin and more or less transparent to translucent. By means of serial microscopical sections bundles of smooth muscle fibers THE SACRO-UTERINE LIGAMENTS 9 have been traced from the uterus toward the rectum, in the recto-vaginal folds. These delicate muscular fasciculi are con- tinuous with the noticeable longitudinal fibers on the dorso- lateral facies of the uterus. It is important to note that the stratum fibrosum of the peritoneum shows marked thickening. The few muscular fasciculi that do reach the sides of the rectum may be traced along its lateral wall to be inserted into its fib- rous sheath, or continuing dorsally, become lost in the meso- rectum. Many of the fasciculi diminish in size as they pass through the folds, their fibers apparently becoming inserted into the peritoneum, but their actual termination has not been observed. Between the two peritoneal layers of the recto- vaginal folds there is a small amount of extremely fine areolar tissue with a few fibro-elastic filaments. In the parametrial and paravaginal tissue there is a very meager amount of fibro-elastic elements. The cephalic extremity of the vagina is freely moy- able and lies between the bases of the broad ligaments. The recto-vaginal folds varied quite a good deal in size and sym- metry in the different guinea pigs examined. 2. BELGIAN HARE Distinct recto-vaginal folds are present. These and the broad ligaments are thin and more or less transparent, containing a varying amount of adipose tissue. Visible fibers are very sparse in the parametrial and paravaginal tissue. The free margins of the recto-vaginal folds are slightly thickened and opaque, as in the guinea pig. This opacity is due to temporary aggregation of the muscular fasciculi. The stratum fibrosum of the perito- neum constituting the folds, shows distinct thickening and at its deep surface passes insensibly into the extremely delicate areo- lar tissue between the layers, where also a few fibro-elastic bundles are found. In the parametrial and paravaginal tissue, there is a varying paucity of fibro-elastic bundles, but no fibers that could be taken to constitute potential ligaments, or no- ticeable aggregations of fibers to constitute true ligaments are present (fig. 5). The fasciculi of the recto-uterine muscle vary 10 FRANK E. BLAISDELL, SR. Reclum_ Plica recto-vaginalis dextra Plica. recto-vaginalis sinistra Excavatio recto-vaginalis Vagina ( post ~partum) Fig. 4 Drawing showing rectum, vagina (post-partum) and recto-vaginal folds in Belgian hare (Lepus europeaus). fasciculi musculi recto-uterin s 7 ‘e! > n f/—feritoneum a 4 om 4 Medial surface Fig. 5 Transverse section of a lateral recto-vaginal fold at middle third, showing distribution of fasciculi of recto-uterine muscle. Belgian hare. X,62. Reduced to one-third THE SACRO-UTERINE LIGAMENTS 11 greatly in size and are distributed chiefly in the apical and lat- eral peritoneal layers of each fold. Many of the fasciculi ap- pear to be almost submesothelial, others which are in the minority appear to be on the inner border-line of the stratum fibrosum, the majority however are in the more central parts of that layer. The muscular fasciculi are abundant in the vaginal ex- tremity of the recto-vaginal fold, but become greatly diminished or absent in the rectal extremity of the same. A large percent- age of the fasciculi therefore terminate in the fold probably by insertion into the more superficial part of the stratum fibrosum. Their actual termination has not been observed. Excavatio re clo-vaginalis Vaginal vault Plica recte-vaginalis Position ef cervix uteri Upson Ligamentum latum YW Cornu uteri — a See i | Fig. 6 Rectum, vagina and uterus of cat (Felis domestica), showing recto- vaginal folds. 3. CAT The recto-vaginal folds are thicker and relatively smaller than in the hare, and pass more noticeably upon the ventral sur- face of the rectum; they are transparent as are also the broad ligaments. The recto-uterine muscles form distinct bands and attain the sides of the rectum; they are fusiform in transverse section and each carries its own blood-vessels. The stratum fibrosum of the peritoneum is distinctly thickened and contains the muscular fasciculi. The fasciculi are most abundant and 12 FRANK E. BLAISDELL, SR. largest in the vaginal extremity of each fold, but many reach the side of the rectum and continue upon the same for a varying distance. They are not all in the prominent part of the fold on the rectum, for as the peritoneum spreads out the fasciculi neces- sarily go with it. The recto-vaginal folds vary in prominence and the distance to which they extend cephalad on the rectum depends on the degree of distention of the latter. It has Plica recto-vaginalis, Fascicult /* ae es eas] Fy musculi recto-uterini/ & —\Muscularis recti Peritoneurn os ™ oe % 7 Fig. 7 Semi-diagrammatic drawing of lateral wall of rectum of cat, show- ing peritoneum and fasciculi of recto-uterine muscle, recto-vaginal folds and muscularis recti (Transverse section). >< 175. Reduced to one-third. been observed that the peritoneum is anchored to the rectum by fibrous trabeculae which arise in the intermuscular fibrous tissue of the muscularis, which by passing outward, through in- tervals in the longitudinal layer become inserted into the stra- tum fibrosum. igure 8 shows the details of a muscular fasciculus, and its more or less centrally located blood vessel. In the majority of cases the vessels are intra-fascicular, but they vary in position to the periphery and may be supra-fascicular. THE SACRO-UTERINE LIGAMENTS 13 4. DOG In the dog, the anatomical relations between the pelvic vis- cera differ considerably from those in the series described above. The vagina during the inter-estral period is rather firmly fixed at its cephalic extremity. The bladder has a short but dis- tinct neck, the peritoneum being reflected from the cervix vesicae upon the anterior vaginal wall before attaining the uterus. In the individuals examined small single or double recto-vaginal folds passed from the utero-vaginal junction to the rectum. The +. it pee ° we eee ®t AS 224; ‘.' sys ro 2 oe Fig. 8 Detailed camera lucida drawing of a fasciculus of recto-uterine muscle with its blood vessel, showing relation to peritoneum (Transverse sec- tion). Cat. > 235. Reduced one-half. ligaments of the bladder and uterus are thin, transparent and contain a varying amount of adipose tissue. The bladder pos- sesses well developed dorso-lateral ligaments which suspend it from the dorsal pelvic wall. The uterus is held in close relation to the ventral surface of the rectum, by its relatively large broad ligaments which pass from the sides of the cervix and cornua, each crossing the dorso-lateral ligament of the bladder of its own side. The recto-uterine muscles pass nearly directly be- tween the two organs, and in the small single or bilateral recto- vaginal folds; or when one or both are absent by way of the peritoneum at the bottom of the recto-vaginal fossa. There is a broad recto-uterine space. Between the folds of the broad liga- 14 FRANK E. BLAISDELL, SR. ments and in the parametrium there is relatively a little more fibro-elastic tissue, but no aggregations which might be con- strued as constituting true or potential ligaments. 5. GENERAL CONSIDERATIONS From the study of the above comparative series the following facts are to be noted. Distinct recto-vaginal folds pass from the dorsal wall and cephalic extremity of the vagina to the ree- tum, the former being more or less free and movable; that these Lipamentum dorso-Jateralis vesica Rectum S| Uterus Plica Distended bladde Vagina. Fig. 9 Drawing of sagittal section of pelvis of a bitch showing relations of viscera and peritoneal folds. recto-vaginal folds vary in size, and that they can be increased by traction on the uterus or more so by traction on both uterus and rectum. The stratum fibrosum of the peritoneum has in all cases undergone thickening or hypertrophy, and the fasciculi of the recto-uterine muscle are contained in it. The fasciculi of the latter are most abundant at the vaginal extremity of the fold and diminish in size and number as the rectum is approached, in some cases not reaching it. Some of the fibers of the muscular fasciculi are apparently inserted into the stratum fibrosum of the peritoneum, fewer of the bundles possibly being inserted into the fibrous tunic of the rectum or lost in the mesorectum. THE SACRO-UTERINE LIGAMENTS 15 The fibro-elastic tissue constituting the parametrial, para- vaginal and paraplical tissue is very scanty in amount and does not form aggregations to constitute so-called true ligaments, nor in sufficient quantity to form potential ligaments. The body of the vagina is more firmly attached than the cephalic extremity. VI. PRIMATE SERIES 1. MONKEY (MACACUS) In the monkey, the uterus is undivided and the anatomical conditions are similar to those in the human female. The weight of the uterus is localized and therefore calls for stronger me- chanical support. ‘There is marked increase in the amount of the parametrial, paravaginal and paraplical fibro-elastic tissue, which can be recognized as constituting potential ligamentous aggregations. The specimens examined had unfortunately been embalmed in the usual way, but nevertheless the material dem- onstrated that the structure and relations were similar to those in the human female. Recto-uterine or sacro-uterine folds were absent, although potentially present. A short plica was pres- ent on each side at the utero-vaginal junction. These folds are not always present in woman, but can always be demonstrated as being potentially present, for traction forward on the uterus brings them into prominence. The thickened peritoneum with the recto-uterine muscles are always present. The histological examination of the material from the monkey was not satisfactory as to details, on account of the amount of cytolysis and softening of the tissues generally, but everything indicated similar structural conditions as in woman. 2. HUMAN For the purpose of establishing a point of departure, the pelvic structures in a state of full maturity or development in woman, will first be taken up, followed by those before maturity and, lastly, those in the stage of senility. [a ae 16 FRANK E. BLAISDELL, SR. a. Examination of the structures in a recent post mortem state The careful dissection of the pelvic viscera of a woman 35 years old gave the following results: The recto-uterine folds were moderate in prominence and diminished gradually, to be- come lost in the general plane of the peritoneum without at- taining the posterior pelvic wall. The rectum was median in position and without a mesorectum opposite the third sacral vertebra. Rectum Ligaonentum i tec ro-uterimiin ey ay al Y/Y a fe) =a - Dae Ligamentum | : c=, Tranisversum Celli) ViuSCUILIS recto-ulerinus Ligamentum pli vaginale-—_— arr Musculus Vaginal vault Fig. 10 Semi-diagrammatic drawing of female pelvic viscera viewed from above, uterus and adnexa having been drawn strongly forward. Position of potential ligaments are shown in black lines. Traction forward on each sacro-uterine fold separately, tight- ened the peritoneum and raised its surface into a series of small ridges, that radiated toward the sacro-iliac junction and lat- eral border of the promontory, from the point where the fold dis- appeared, in a direction downward and inward to the middle of the second sacral vertebra at the edge of the rectum and reflec- tion of the peritoneum upon it. The area raised on the right was triangular but did not extend so far laterally as given in figure 10; on the left side the raised area was narrower and less triangular than on the right. Traction backward on each fold THE SACRO-UTERINE LIGAMENTS Life tightened the peritoneum about the uterine cervix and vaginal vault, raised the cervix upward and backward, forced the fundus against the bladder and raised the vaginal vault. Traction on the uterus to the right or left simply tightened the broad ligaments, peritoneum, and subperitoneal fibro- elastic tissue without any special result. Bodies embalmed in the usual way for dissection, or with a 10 per cent to a 40 per cent formaldehyde solution, will usually show the same result of traction on the sacro-uterine or recto- uterine folds, and parametrial fibro-elastic tissue as above stated. The embalming fluids used, especially the formaldehyde solution, produced marked contraction of fibro-elastic tissue and such contraction produces bands which are rendered slightly prominent as peritoneal ridges, more or less above the general surface contour of the peritoneum, the results varying in dif- ferent cadavers. Two female cadavers in the anatomical col- lection of the university show these facts beyond doubt. Traction per vaginum downward and forward on the uterus depresses the sacro-uterine or recto-uterine folds and draws on the sacral attachments of the peritoneum, in the line of the folds. At the same time there is a coincident appearance of a prominent ridge in each dorso-lateral wall of the vaginal vault, corresponding to the position of the sacro-uterine and _plico- vaginal ligaments. There is a tightening of the subperitoneal tissue laterally as well as dorso-laterally to the uterus. Reflection of the peritoneum lateral to the uterus and recto- uterine fold. uncovers a mass of fibro-elastic tissue, through which run the branches of the hypogastric vessels, lymphatics, and nerves on their way to the uterus, vagina and bladder. Traction in different directions on this mass demonstrated its great elasticity as well as the intrinsic movements of the fascic- uli and lamellae over each other. Careful dissection and exam- ination with a moderately strong hand lens clearly defined a mass of interlacing fibers and fasciculi ensheathing the vessels and nerves and having an attachment to the fascia covering the levator ani, coccygeus and obturator muscles, as well as the pre- sacral fasciae and peritoneum. Pulling on the fibro-elastic THE ANATOMICAL RECORD, VOL. 12, No. 1 18 FRANK E. BLAISDELL, SR. tissue from different points laterally and at the periphery, drew the uterus in the same direction; drawing the uterus toward the opposite side of the pelvis tightened the fibro-elastic mass which appears to radiate chiefly from the sides of the uterine cervix, while traction toward the middle of the inguinal ligament tight- ened the fibro-elastic mass laterally and posteriorly on the opposite side. The examination of several autopsy specimens not only veri- fied the above facts but revealed others (fig. 10). By carefully cutting through the peritoneum in the anterior part of the lat- eral wall of the recto-vaginal fossa, dissecting out the areolar tissue filling in the meshes of the fibro-elastie net-work and iso- lating a number of fibers, a dimpling of the peritoneal surface along the sacro-uterine fold could be produced by pulling down- ward on the fibers. Other fibers appeared to be attached fur- ther posteriorly in the vicinity of the sacrum. Pulling up- ward on the same fibers raised the vaginal vault or the cervix uteri. These experiments determined that there were two sets of fibers which entered or passed beneath the fold. One set passing from the sides of the uterine cervix backward, giving off fibers which were inserted into the stratum fibrosum of the peritoneum along the line of the fold, and others reaching the presacral fascia. The second set passed from the sides of the vaginal vault below the above and inserted into the stratum fibrosum along the anterior two-thirds of the fold. The study also determined that the meshes of the fibro-elastic network varied in size and irregularity. At times there was observed an — apparent increase in the density of the fibro-elastic mass opposite to the cervix uteri and vaginal vault. At the periphery of the pelvic cavity just in front of the sacro-iliac articulation, the network appeared less dense. This condition was not always evident. In cadavers that have been embalmed in the usual way and dissected, there will be found fasciculi passing laterally from the side of the uterine cervix, and forming a more or less dis- tinet band which corresponds in position to the cardinal ligament or ligamentum transversale colli. Other fasciculi which pass THE SACRO-UTERINE LIGAMENTS 19 backward beneath the sacro-uterine fold to reach the presacral fascia, correspond to the course of the so-called sacro-uterine ligament. The fibers sweeping downward from within the an- terior two-thirds of the sacro-uterine fold to be inserted into the sides of the vaginal vault, are termed the ‘plico-vaginal ligament.’ b. Histological observations. Fixed material 1. Adult 35 years old. Figure 11 is a photomicrograph of a portion of a transverse section of the sacro-uterine fold of a 4 Peritoneum Plica sacro-uterinae Vessels Fibro-elastic fasciculi and lamelle Fig. 11 Photomicrograph of a transverse section of recto-uterine fold at middle of anterior third, of a woman 35 years old. X 50, oc. 2; obj. A. woman 35 years old. The section is through the anterior third of the fold and includes the vaginal vault. The stratum fibro- sum of the peritoneum is thickened and dense, the muscular fasciculi are abundant within it. 20 FRANK E. BLAISDELL, SR. The inner face of the dense fibrous layer shows a distinct ten- dency to break up into segments or divisions and most of them ap- pear to be directly continuous with the fibro-elastic fasciculi or lamellae which sweep downward out of the fold to become part and parcel with the general fibro-elastic and perivascular net- work. Beneath the peritoneum therefore, the fibro-elastic Subperitoneal ; eee Sauk Muscular fascicult Fig. 12 Photomicrograph, transverse section of right recto-uterine fold in anterior part of middle third, of a child 10 years old. X 30, oc. 4; obj. A? (Zeiss). tissue is dense and rich in blood vessels, lymph vessels and nerves. ‘These fibro-elastie fasciculi and lamellae are parts of the plico-vaginal and sacro-uterine ligaments. The _ plico- vaginal constituents end in the inner surface of the thickened peritoneum, and are the fibers which caused the dimpling in of peritoneal surface of the fold in the experiments reported above. THE SACRO-UTERINE LIGAMENTS “| 2. Child 10 years old. In sections (fig. 12) made at the middle of the sacro-uterine folds the peritoneum is thickened as usual and small fasciculi of muscle fibers are scattered sparsely through the slightly dense stratum fibrosum. On the border line be- tween the latter and the subperitoneal tissue there is an aggre- gation of numerous large muscular fasciculi, small vessels and nerves. These fasciculi are surrounded by rather loose fibro- elastic tissue, but the general character of it is such that it can be associated with the peritoneum, rather than with the sup- peritoneal tissue. In ell probability, as growth continued, a Rectum E ‘: __Plica vecto-uterina sinistra xCavatio recto-uterina _4 Vaginal vault — . 4% Cystic ovary Ut Cystic ovary se Fig. 13 Drawing showing the female pelvic viscera in an infant 40 days old. The uterus has been drawn forward. Made from the specimen studied. greater condensation would take place and these elements would become more closely bound to the peritoneum. The sections are from behind the plico-vaginal ligament, and there are no fibers sweeping downward which could be taken for it. It is possible that a part of the above mentioned muscular fasciculi are those that accompany the sacro-uterine ligament. 3. Infant 40 days old. Figure 13 is a drawing of the pelvic viscera of an infant 40 days old. The specimen was cut in serial transverse sections. The left sacro-uterine fold was very promi- nent and terminated lateral and dorsal to the rectum. The right fold was much less prominent and terminated upon the lateral 22 FRANK E. BLAISDELL, SR. -all of the rectum. The structure of the folds as regards den- sity of the tissues, was similar to those of the adult 35 years old. F igure 14 is a photomicrograph of a section through the middle of the right fold, the peritoneum is thickened, and in the flattened apex of the fold the intraplical fasciculi are very inti- mately connected to the deep surface of the stratum fibrosum; Plico-vaginal fasciculi mellae and lamellar Fig. 14 Photomicrograph of a transverse section through the middle of the right recto-uterine fold, of an infant 40 days old. Note the dimpling of the peritoneal surface. > 50, oc. 4; obj. A. in the latter there are small muscular fasciculi. From their points of insertion into the peritoneum, the fibro-elastic fasciculi stream downward lateral to the recto-uterine fossa and become continuous with the general perivascular tissue. The photomicrograph shows the wrinkling of the peritoneal surface and the fibro-elastie fasciculi can be definitely traced downward from their attachment to the stratum fibrosum. In THE SACRO-UTERINE LIGAMENTS 23 both folds, these fasciculi are plico-vaginal and sacrc-uterine, the former being the most evident. The intraplical muscular fasciculi are those which in part probably accompany the sacro-uterine fibro-elastic fasciculi. To what extent these muscle bundles become associated with the peritoneum to constitute recto-uterine muscle bundles is not known. Fig. 15 Photomicrograph of a transverse section of a right plica recto- uterina at anterior third, through plico-vaginal ligament (a), showing peri- vascular (b) character of fibrous tissue, meshes (c) between the lamellae (d) and fasciculi (e) filled with delicate areolar or adipose tissue (the former has not been filled in); lateral to ligament the tissue is distinctly fibro-elastic and areolar (f). Infant 40 days old. X 88 diam., oc. 2; obj. A. Reduced to one-third. Figure 15 is a drawing of the paraplical tissue below the point shown in figure 14. It shows the perivascular character of the fibro-elastic fasciculi and lamellae. The tissue shown here is a part of the plico-vaginal ligament. Particular atten- tion should be given te the manner in which the mesh-work is formed—by division and union of fasciculi and lamellae and 24 FRANK E. BLAISDELL, SR. how they enclose the vessels. The whole is representative of the arrangement throughout the parametrial, paravaginal and para- plical network. It has been observed that the lymphoglandulae are not as a rule enveloped by the fibro-elastic lamellae, but pro- ject into a mesh, being only attached at the hilus to a lamella, which immediately envelops the vessels entering or leaving it. At least this has been the case in a number of instances. The capsule of the gland is continuous with the lamella at the hilus. Elastic plate Museular Atrophic tissue fasciculi Peritoneum Fig. 16 Photomicrograph of a transverse section of a sacro-uterine fold at the posterior third, of a woman 65 years old. X 50, oc. 2; obj. A. The meshes of the network are filled with delicate areolar or adipose tissue, as shown in figure 15, where a part of the meshes have not been filled in, to better demonstrate the fibro-elastic network. Lateral to the plico-vaginal fasciculi, the tissue is distinctly elaStic and areolar. 4. Adult 65 years old. Figure 16 is a photomicrograph of a portion of a transverse section of a plica sacro-uterina of a woman THE SACRO-UTERINE LIGAMENTS 25 past the menopause. The peritoneum still remains much thick- ened, the bundles of the recto-uterine muscle are very distinct, and the subperitoneal fibro-elastic tissue shows distinct senile atrophy. The general looseness of the tissue is noticeable and adipose tissue is more abundant than at the other ages con- sidered above. Figure 17 is a drawing of a portion of the peritoneum in the section from which figure 16 was made, but more highly magni- fied. The muscle bundles have been drawn in heavy black to CUPS ade Elastic plate fasciculi musculi recto-uterini Z Per oe. Subperitoneal eritoneum ___ eee, Ou bperitonea Ys ie tissue Fig. 17 Photomicrograph of part of a transverse section of plica sacro- uterina at posterior third, showing distribution of fasciculi of recto-uterine muscle, elastic plate and subperitoneal connective tissue in a woman 65 years old. X 88. Reduced to one-third. emphasize their abundance. At the inner limit of the dense stratum fibrosum there is a distinct layer or plate of elastic tissue. Figure 18A shows this layer as brought out by Weigert’s elastic tissue stain. This elastic plate lies to the inner side of the thick- ened fibrous stratum of the peritoneum, and becomes much thinner at the periphery of the plical area where it approaches quite close to the mesothelium; it appears to limit internally the stratum fibrosum. Whether or not this layer will be of value in 26 FRANK E. BLAISDELL, SR. determining the relation of the recto-uterine muscle to the peri- toneum remains to be seen. Figure 18B illustrates the abun- dance of elastic fibers in a fasciculus of the recto-uterine muscle of the same section. 5. Adult 83 years old. Figure 19 shows one of a transverse series of sections through a sacro-uterine fold of a woman 83 years old. The section has been taken from the fold at the middle of the middle third. The fibrovs stratum of the peri- toneum is thickened as usual, and immediately beneath the mesothelium there is a distinct layer of undulating fibers (A), that separates the stratum containing the muscular fascic- uli from the mesothelium. The fibers of the deeper layer (B) are more irregular and broken, receiving the insertion of the Fig. 18 A. Section of peritoneum from sacro-uterine fold of a woman 65 years old. B. Transverse section of a recto-uterine muscle, bundle, showing abun- dance of elastic tissue. Weigert’s elastic tissue stain. X 90 diam. Camera lucida drawing. Reduced one-half. plico-vaginal fibers which are well shown in the sections. The fibro-elastic tissue is less compact than in the infant 40 days or the woman 35 years old on account of the senile atrophy present. Weigert’s elastic tissue stain also brought out a greater irregu- larity in the distribution of the elastic fibers. The elastic plate so well defined in figure 17 is missing here, although a very irregular line of elastic fibers is present. The stain has also brought out a greater amount of elastic tissue in the submesothe- lial fibrous layer that was not noticeable in the sections from which figure 16 was made. In figure 18B there is shown an abundance of elastic fibers in relation with the muscular fascic- uli and it is to be noted that they are between the elastic plate THE SACRO-UTERINE LIGAMENTS 2 Sacro-uterine lamella Peritoneum Containi ng Muscular fasciculi Plico -vaginal fascicull Fig. 19 Photomicrograph of a transverse section of sacro-uterine fold at middle of middle third, in a woman 83 years old. X 30, oc. 2; obj. a? (Zeiss). and the mesothelium. Beyond the area of distribution of the fasciculi of the plico-vaginal and sacro-uterine ligaments, adi- pose tissue is very abundant, scattered through which is a vary- ing but meager number of fibro-elastic lamellae that have been cut across. VII. VARIATIONS It is well known from observations made in the dissecting room and at autopsies, that the peritoneal surface contour of the pelvic cavity and relative size of the pelvic viscera are sub- ject to considerable variation; at times to marked asymmetry. The bones forming the pelvic wall may vary so as to render the cavity narrow and relatively deep, or broad and relatively 28 FRANK E. BLAISDELL, SR. shallow. The muscles may be much better developed in some individuals than in others, and the amount of fibro-elastic tissue may likewise vary. Every gynecologist knows that the perineal body may be large and strongly constituted in some women, or feeble and scarcely recognizable. The muscles guarding the pelvic outlet are very strong and capable of affording ample support to the pelvic organs in some individuals, in others they are poorly developed and incapable of withstanding continued strain. These variations bespeak for a general habitus which may indi- ‘ate a predisposition to the persistent maintenance of the norm, or an easy deviation from the same. The plicae sacro-uterinae are no exception to the rule. In certain cases they are strongly developed and very prominent and symmetrical or asymmetrical; at other times they apparently are entirely absent, or a short plica may be present on each side near the uterus. But even when visibly absent, they are potentially present, for forward traction on the uterus produces them. Or a distended rectum, by carrying the peritoneum away from the pelvic wall, will produce them or convert sacro-uterine into recto-uterine folds. The position of the rectum also influences the character of the folds. If the rectum is median in position, both folds are usu- ally sacro-uterine; if that organ is sinistral, the corresponding fold will be recto-uterine and the opposite will be sacro-uterine and vice versa when the rectum is dextral in position. Histo- logical examination reveals similar variations, both as regards quantity and symmetry, in the uterine supports. VIII. EXPERIMENTS In an anesthetized cat, with the abdomen opened and the pelvic organs exposed to view, the uterus and bladder have been observed to contract more or less intermittently as peristaltic waves passed down the rectum. Though slight, these con- tractions were unmistakable. In the animal studied, the rectum happened to be strongly distended with fecal matter opposite the recto-vaginal fossa and the point of attachment of the recto-vaginal folds. The THE SACRO-UTERINE LIGAMENTS 29 viscus was stretched in the directions of both its primary and secondary axes. The recto-vaginal folds were large and ex- hibited occasional contractions, which feebly raised the vaginal vault. After the sigmoid colon had been severed and the fecal contents expelled, it was observed that the intestine slowly contracted and sank lower in the pelvis. During this descent the recto-uterine folds diminished in size and the vaginal vault sank carrying the uterus with it. The rectum was then injected to reproduce the effect on the vaginal vault through traction on the recto-vaginal folds. The rectum was clamped behind the free extremity of the vagina, and the rectum slowly distended with water. As a consequence, the rectum pulled cephalad on the folds raising the vaginal vault and carrying the uterus toward the abdominal cavity. These observations in the cat suggested, first, that the distended rectum mechanically raises the vaginal vault when its fecal contents are passing caudalward and thus prevents compression of the organs of reproduction; second, the act in all probability excites a reflex contraction in the uterus which participates in the act through the recto-uterine muscles; third, that the uterus can automatically raise itself through con- traction of the recto-uterine muscles in the recto-vaginal folds. These experiments were repeated on a cadaver of an infant 40 days old. The uterus was raised as before by traction on the right recto-uterine fold by distention of the rectum, and at the same time the partial conversion of the left sacro-uterine fold into a recto-uterine fold was accomplished. Hence the uterus can automatically raise and tilt itself ventrad in primates, or be raised mechanically by the recto-uterine folds when the rectum is distended. Or, distention of the rectum may cause reflex contractions of the uterus. When the sacro-uterine folds are present and the rectum is median in position, the uterus must act alone through a reflex. If the rectum is sufficiently distended, it can convert sacro-uterine folds into recto-uterine folds. When the uterus is raised through traction or contrac- tion of the recto-uterine muscle, the sacro-uterine fold is tight- ened, the plico-vaginal ligament is pulled apward carrying the vaginal vault with it. 30 FRANK E. BLAISDELL, SR. IX. THE MUSCULI LEVATORES UTERI No reference has been made to the muscular tissue that passes off laterally from the uterus to be dispersed through the fibro- elastic tissue filling the subperitoneal space lateral to the uterus. Microscopical examination of the parametrial tissue shows that smooth muscle is abundant about the point of insertion of the fibro-elastic tissue into the sides of the uterine cervix, and that it diminishes in abundance as it is traced laterally and poste- riorly. The fibers appear to be inserted into the perivascular tissue, the fibrous fasciculi and fasciae of the nearby muscles, but their actual termination has not been determined. Direct continuity of the recto-uterine muscles with the muscularis uteri is established. This muscular tissue extends from the uterus into the adjacent tissue at an early embryonic period. These muscles, one on each side of the uterus, enable that organ to automatically raise itself on the fibroelastic suspensorium. By their more or less intermittent contractions, they probably become one of the chief agents in aiding the venous circulation in the peri-uterine venous plexuses; the pelvic diaphragm being accessory to the act. X. DISCUSSION In taking up the discussion of the uterine ligaments, it is necessary to consider briefly the mechanical supports of the uterus in order to fully appreciate the part which the fibro- elastic or so-called perivascular tissue plays in the process. The mechanical supports of the uterus will be considered in the following order: 1. The levator ani and its superior fascia. 2. The peritoneum. 4. The fibro-elastic tissue filling in the interval between the two first mentioned. Gynecological text-books usually speak of the pelvic floor as & support of the uterus. Fothergill (15) and Cameron (13), the former preceding the latter, have described the levatores ani as forming ‘a tunnel’ on each side of the vagina and being attached THE SACRO-UTERINE LIGAMENTS 31 to it at its lower part. Cameron (13) is quite right when he states that the uterus which is above the vagina—the latter not receiving any direct support in its upper part from the muscular diaphragm—has no support from these muscles. The superior fascia of the levatores ani does not prevent the de- scent or ascent of the uterus. He asks why it is that the super- incumbent intestines do not crowd the uterus and the vagina downward concertina-like? Cameron (13) rightly concludes that some other structures prevent this, which according to him are the ‘perivascular fascia’ and the blood vessels. It is certain that the pelvic diaphragm plays a large part in supporting the pelvic viscera, and indirectly the descent of the superincumbent intestines, which when normally suspended by their mesenteries exert very moderate pressure. The normal supporting function of the pelvic diaphragm may be likened to a foundation upon which a superstructure rests, and which when it ascends or descends, carries the superstructure with it. By virtue of its contractile power and up and down movement, it also aids in preventing venous stasis within the pelvic plexuses, besides fulfilling other functions. The superior fascia of the levatores ani steadies and aids in maintaining the cephalic extremity of the vagina and uterus in their median position, and also gives support and attachment to the parametrial fibro-elastic mesh-work. The peritoneum may be regarded as a support in the sense that it envelopes the pelvic organs as a sheet, forming lateral folds for the support of vessels and the adnexa uteri, maintaining the uterus in its normal anatomical position, and preserving this relation in its physiological excursions. Besides, as Cameron (13) states, it furnishes the superior attachment of the ‘perivascular tissue.’ The sacro-uterine or recto-uterine fold of the peritoneum has a varying degree of usefulness, which has already been partially stated and will be referred to again. Cameron (13) denies that these folds exert any supporting influence on the uterus, but laid great stress on the ‘perivascular tissue,’ which is weakest an- teriorly lateral to the bladder, and between the folds of the broad ligaments; but increases in thickness as it is traced back- 32 FRANK E. BLAISDELL, SR. ward, and is greatest in amount opposite the broad ligaments and the sacro-uterine folds. Its attachments, to quote Cameron (13) again, are as follows: ‘‘ Above to the peritoneum, below to the pelvic sheaths of the levator ani and coccygeus, externally to the obturator fascia, and higher up to the periosteum of the innominate bone, while internally it blends with the connective tissues of the viscera. Its attachment to the lateral pelvic wall is rendered further secure by the fact that the parietal branches of the hypogastric vessels pierce the pelvic parietes in order to reach their respective destinations, and in doing so their sheaths blend with the surrounding bony and muscular structures.’ It is admitted that the ‘‘main trunks of the hypogastric vessels, in their descent on the lateral wall of the pelvis, are bound down to the latter by dense areolar tissue.” Cameron (13) has somewhat over-estimated the relative size of the vessels passing to the bladder, uterus and vagina. The main vessels are large and well anchored to the lateral wall of the pelvis, and their visceral branches are supported and sur- rounded by the fibro-elastic tissue in part. These vessels are not a factor in the support of the uterus, and the ovarian ves- sels enter too high to be a support; and besides, it is to be ex- pected that vessels which are to be put to greater or less stretch- ing, will be more or less tortuous in their course. The parametrial tissue contains the peri-uterine venous plexuses, which must not be kept on a strain or under continual compression. It is necessary to again call attention to the abundance of the lymphatic vascular net-work, and the numer- ous nerves that permeate this region. The round ligaments are of doubtful importance as uterine supports, and Fothergill (15) has pointed out that they are essentially embryonic in function. Cameron (13) has given importance to the obliterated hypo- gastric arteries as a pelvic support. They may act feebly. They are essentially embryonic in function. Cameron (13), after his discussion of the importance of the different pelvie sup- ports, sums up by stating that ‘‘the perivascular fascia, plus the pelvic sheaths of the levator ani and coceygeus muscles, are the most important.” THE SACRO-UTERINE LIGAMENTS 33 Montgomery (8) considers that the supports of the uterus are not ligaments in the ordinary sense, but consist of connective tissue, into and through which run prolongations from the uterine muscular structure, so that the organ is virtually sus- tained by muscular action. But he confounds the movements of the pelvic diaphragm with true intrinsic uterine activity, such as it is capable of exerting through the musculi levatores uteri. Its excursions upward and downward with ‘every respira- tory’ movement, depends upon the action of the respiratory mechanism. Mackenrodt (3) lays great stress on a band of connective tissue, the ligamentum tranversale colli, as of great physiologi- cal importance in maintaining the normal position of the uterus. He recognizes that the lower opening of the pelvis is closed by pelvic fascia, which sends firm bands to the cervix and vagina; that the cervix is held fast in its embryological position by liga- ments, while the uterine body is kept in position by its own weight and intra-abdominal pressure—not by ligaments. Evi- dently this knowledge of the ‘firm bands’ was acquired through the dissection of embalmed material. He says that fibers com- ing from the pelvic fascia to the side of the cervix are to be sharply defined from the sparse connective tissue between the folds of the broad ligament. These, he states, form a band that is the chief means of holding the uterus in position. Another statement is that this band or ligementum trensversale colli carries the arteria uterina in its upper part (vide fig. 20). Ovenden (12), in her dissections, found very “‘little connective tissue between the layers of the broad ligament, but at the level of the cervix, however, a thick band can be felt between the two layers of the peritoneum. It is wedge-shaped in section; the apex of the wedge is directed upward and is just above the level of the point of entrance of the uterine artery. Traced to its distal attachments, this band is found to be formed from strong fibrous connective tissue, continuous with that which surrounds the pelvic blood vessels, and also that which comes through the sacro-sciatic notch. Some of the fibers appear THE ANATOMICAL RECORD, VOL. 12, No. 1 34 FRANK E. BLAISDELL, SR. also to be attached to the sides of the third and fourth pieces of the sacrum.”’ Mackenrodt (3) considered that the ligamentum transversale colli has its central attachment at the supra-vaginal portion of the cervix. Ovenden (12) considers it inserted partly into the vaginal vault and lateral fornix, besides into the sides of the uterus for a short distance below the point of entrance of the uterine artery (vide figure 20). Tuba uterina. Lig. tetes uteri. Art. ovarice. Peritoneum. Art. ulerina a aN Lig. sacrouterina. Mucosa vagme Fig. 20 Side view of uterus (as removed by vaginal hysterectomy) showing insertion of ligamentum transversale colli of Mackenrodt (after Ovenden). These facts agree with the observations reported in the ear- lier part of this paper. Ovenden (12) is correct in considering that the whole mass is not inserted into the uterus, as Macken- rodt (3) asserts; and also that the sacro-uterine ligament blends with the ligamentum transversale colli near its insertion into the uterus. The plico-vaginal ligament attached to the vaginal vault becomes continuous with the sacro-uterine ligament. It is to be noted that these observations make these three liga- ments continuous at their insertion into the sides of the cervix and vaginal vault. THE SACRO-UTERINE LIGAMENTS 390 It must be mentioned that Emmet (23) and Schauta (24) have laid emphasis on the importance of the part played by this pelvic connective tissue in maintaining the normal position of the uterus, although they did not ascribe this function to a particu- lar band. Specimens from the cut ends of the so-called sacro-uterine ligaments, as severed from their attachment into the sides of the uterine cervix at the time of operation for utero-vaginal pro- lapse in elderly women, were submitted to the writer by Dr. George B. Somers (25) for microscopical study. One ligament, evidently cut closely to the uterus, showed a great preponder- ance of smooth muscular tissue over the fibro-elastic; the other, much less muscular and chiefly fibro-elastic tissue. Ovenden (12) states that in microscopical sections the ligament consists largely of fibrous tissue, through which are scattered a good many bundles of smooth muscle fibers. None of the writers have attempted to explain the marked elasticity that is inherent in the pelvic structures, not how or why the uterus can be pulled down to the introitus vulvae or to the exterior, and when released will slowly and completely re- turn to its normal position in the pelvic cavity without further manipulation. From the experiments, dissections and observa- tions reported in this paper, it now becomes necessary to de- scribe the fibro-elastic suspensorium uteri, which accounts for all of the phenomena that have thus far been observed and reported. If a square piece of thin paper be taken and folded over two or three times, cut one-half across first on one side and then on the other, when opened up it will appear as in figure 214A. Traction applied in direction of the arrows, or diagonally at the corners, will open up a mesh-work as in figure 21B. Release the paper and it will instantly return to a state of rest as before the traction was applied (fig. 214A). This paper possesses elasticity and an inherent tendency to return to a state of rest, but the phenomena are in a reverse order to that observed in the fibro- elastic mesh-work of the suspensorium uteri. 36 FRANK E. BLAISDELL, SR. Figure 22A represents the fibro-elastic mesh-work filling the parametrial and paraplical space in a state of physiological tonus or rest (potential ligament). It is an open mesh-work, P | Fig. 21 A piece of tissue paper folded and cut (A) so as to forma mesh work (B) that possesses elasticity and an inherent tendency to return to a state of rest as in A, but in a reversed order to that of the fibro-elastic mesh-work of the suspensorium uteri. | Ni He Aiea Cle f= ne NN / WANS Fig. 22 Drawings illustrating the effect of traction upon fibro-elastic mesh- work. (A) Fibro-elastic mesh-work in state of physiological tonus or rest (potential ligament); (B) fibro-elastic mesh-work in state of traction (actual ligament). THE SACRO-UTERINE LIGAMENTS etl the meshes of which are filled with areolar or adipose tissue. If traction is made as indicated by the arrows, the condition seen in figure 22B will result, the fasciculi and lamellae will be approximated and a transitory ligament will form. Remove the traction and by virtue of the inherent elasticity or resiliency the mesh-work returns. These simple experiments demonstrate the mechanism of the fibro-elastic suspensorium uteri. The fibro-elastic network is like a net that has been sym- metrically attached at a periphery and inserted into the two sides of a body, which is suspended by it. Traction downward on this body approximates the threads of the net which appear to diverge from the point where they are attached to it. If the body is released it is immediately carried upward until the in- herent elasticity of the netting is satisfied, when a state of rest is established. The threads of the net no longer appear diver- gent at the sides of the body for they appear as a part of the net-work. Such is the author’s conception of the manner in which the uterus is suspended by the fibro-elastic tissue of the parametrial and paraplical space and which is aided by the pelvic diaphragm and peritoneum. From what has already been said in this paper, the full import of the argument should be clear to all. The normal workings of the suspensory mechanism is best observed in the virgin pelvis. Various factors begin to operate at the time of puberty, which may sooner or later weaken the power of the uterine or vaginal supports. Aside from the weakening effects of parturition, with possible injury to the pel- vie floor, the pernicious effects of chronic constipation and tight lacing have to be taken into account. These slowly cause the fibro-elastic tissue to give away with loss of the normal resil- lency or tone. The writer agrees with Fothergill (15) in stating that pro- lapsus uteri may not occur even in long standing laceration of the perineum, providing the fibro-elastic suspensorium retains its tone. In the section on variation, the writer has pointed out that the relative size and strength of parts may vary greatly in the same and different individuals. A weak pelvic floor will 38 FRANK E. BLAISDELL, SR. not bring about prolapsus uteri if there happens to be a well developed fibro-elastic suspensorium; on the other hand, a strong pelvic floor will delay a prolapse when 2 weak suspen- sorium is present. A lacerated perineum is always a source of danger. The applied facts should be clear. With Cameron (13), the writer suggests that an attempt must be made to restore the weakened fibro-elastic support, if success in treatment of prolapse and mal-positions of the pelvic organs is to be obtained. XI. CONCLUSIONS The material studied for the preparation of this paper seems to justify the following deductions and problems: The plicae sacro-uterinae or recto-uterinae are the homo- logues and analogues of the plicae recto-vaginales of quadrupeds. The musculi recto-uterini of the higher Primates are the homologues and analogues of the same in quadrupeds. The plicae recto-uterinae and their intimately associated musculi recto-uterini 2re primitive, appearing in the vertebrate series before the fibro-elastic suspensorium uteri or a well devel- oped musculus levator uteri. The suspensorium fibro-elasticum uteri has been gradually evolved with the assumption of the erect attitude in locomotion, and is not present, or is present only in a very rudimentary way in animals which habitually assume the horizontal attitude in locomotion. If present in a rudimentary or primitive form, it is only concerned in maintaining to a greater or less extent the cephalic portion of the vagina. In woman, and the females of the higher Primates at least, the supports of the uterus are three in number, namely: 1. The suspensorium diaphragmaticum (pelvis). . The suspensorium peritoneale. . The suspensorium fibro-elasticum. The latter being the chief and essential support, the others being accessory. The suspensorium fibro-elasticum consists of a fibro-elastic network supporting vessels and nerves, and contains the poten- 2 2. v THE SACRO-UTERINE LIGAMENTS 39 tial ligaments of the uterus and vagina. The meshes of the network are filled with aerolar tissue which permit the fasciculi and lamellae to move freely over each other. The potential ligaments are the ligamentum transversale colli of Mackenrodt, ligamentum sacro-uterinum, and the liga- mentum plico-vaginale, which become actual through traction on the uterus and vaginal vault. The three above-named ligaments are directly continuous with each other at the sides of the uterine cervix and vaginal vault and with the general fibro-elastic network to the periphery. The fibro-elastic network in a state of physiological rest forms an open mesh-work and possesses an inherent tendency to return to a state of rest after traction on the uterus or va- ginal vault has ceased to operate, and neutralizes or minimizes downward pressure through this same property. The fibro-elastic network in a state of rest prevents com- pression of the venous and lymphatic vessels by preventing collapse of their walls. All traction upon it more or less com- presses the vessels and it hence becomes an aid to the venous and lymphatic circulations. The upward and downward movements of the suspensorium diaphragmaticum augments the action of the suspensorium fibro-elasticum in aiding the pelvic circulation, and in this way it is analogous to the thoracic diaphragm. The suspensorium fibro-elasticum permits the uterus being depressed to the introitus vulvae. The return of the uterus to its normal position being first aided by the diaphragmatic funnel, and completed by the suspensorium fibro-elasticum. The musculi levatores uteri are derived trom the muscularis uteri and constitute a mechanism by which the uterus can automatically raise itself on the suspensorium fibro-elasticum. By its more or less rhythmical or intermittent contractions, in conjunction with the muscularis uteri, it aids the venous and lymphatic circulation in the respective peri-uterine plexuses. The plicae sacro-uterinae or recto-uterinae, with the in- timately associated musculi recto-uterini, lifts the uterine 40 FRANK E. BLAISDELL, SR. cervix and vaginal vault upward and backward, through stimuli received from a distended upper rectum or from other sources. Those fasciculi and lamellae of the fibro-elastic network that arise from the presacral fasciae and along a plica sacro-uterina, and which have a general trend to the vaginal fornix and uter- ine cervix, behind and below the point of attachment of the ligmentum transversale colli, constitute the ligamentum sacro- uterinum, potential or actual (transitory). Those fasciculi and lamellae of the fibro-elastic network arising from the fasciae of the levator ani, obturator, or sheath of the hypogastric vessels in the lateral wall of the pelvis, and with a general trend toward, and attachment to the side of the uterine cervix below the uterine artery, and above the ligamentum sacro-uterinum, constitute the ligamentum trans- versale colli potential or actual (transitory). Those fasciculi and smaller lamellae arising from the stratum fibrosum of the peritoneum of the anterior two-thirds of a plica sacro-uterinum or recto-uterinum, and inserting into the sides of the vaginal vault below the ligamentum sacro-uterinum constitute the ligamentum plico-vaginale, potential or actual (transitory). Any weakening of the suspensorium diaphragmaticum, which will result in a falling of the pelvie floor, or when coupled with a lacerated perineum, will result sooner or later in an over- stretching, and a giving way of the suspensorium fibro-elasticum, and must result in a vaginal or uterine prolapse, or malposi- tion of the uterus, with consequent disturbance of the fibro- elastic and muscular mechanisms, which will be rendered more or less inoperable, with resulting venous stasis and increased weight of the pelvic viscera. In conclusion, the writer desires to acknowledge his indebted- ness to Prof. A. W. Meyer, for advice, and to Prof. William Ophils and Dr. Edgar D. Downing, for the permission to examine and use material from the autopsy room; to Prof. E. ©. Dickson for the preparation of the photomicrographs. The work was begun while the author was connected with the division of anatomy. THE SACRO-UTERINE LIGAMENTS 41 XI. LITERATURE CITED (1) Kocxs, J. 1880 Die Normale und Pathologische Lage und Gestalt des Uterus. Bonn. (2) Hart, J. Berry 1880 The structural anatomy of the female pelvic floor. Edinburgh. (3) MacxenropT, A. 1895 Ueber die Ursachen der normalen und patholog- ischen Lagen des Uterus. Archiv f. Gyn., 48, p. 393. (4) Hott, M. 1897 Die Muskeln und Fascien des Beckenausganges. Von Bardelebens Handbuch der Anatomie des Menschen, Bd. 7, Jena. (5) Harman, N. Bisoop 1898 The pelvic splanchnic nerves. Journ. Anat. and Physiol., vol. 33, p. 386. (6) THompson, P. 1900 The pelvic diaphragm. Studies from the Anat. Dept. of the Owens College, vol. 2. (7) Deaver, JonHn B. 1903 Surgical Anatomy, vol. 3. (8) Monrcomery, E. E. 1903 Practical Gynecology. (9) Stony, R. A. 1904 The anatomy of the visceral pelvic fascia. Journ. Anat. and Physiol., vol. 38. (10) Derry, Dovetas E. 1907 On the real nature of the so-called pelvic fascia. Journ. Anat. and Physiol., vol. 42, Ist Pt., Oct. (11) Paterson, A. M. 1906 The mechanical supports of the pelvic viscera. Brit. Med. Journ., Dee. 15, p. 1701. 1906-07 The mechanical supports of the pelvic viscera. Journ. Anat. and Physiol., London, 41, p. 93. (12) OveNDEN, Exta G. 1906-07 The lateral fixation of the cervix uteri. Jour. Anat. and Physiol., London, 41, p. 308. (13) Cameron, JoHN 1907-08 The fascia of the perinaeum and pelvis of the female, with special reference to the mechanical supports of the pelvic viscera. Journ. Anat. and Physiol., 42, Ser. 3, p. 438. (14) Smita, G. Etuiorr 1908 Studies in the anatomy of the pelvis with special reference to the fasciae and visceral supports. Pt. 1, Jour. Anat. and Physiol., Jan., p. 198. (15) ForHeraitt, W. E. 1908 The supports of the pelvic viscera. A review of some recent contributions to pelvic anatomy with a clinical intro- duction. Jour. of Obstet. and Gyn. of Brit. Emp., Jan. (16) Somers, Gro. B. AND BLAISDELL, F. E. 1913 The anatomy and surgical utility of the sacro-uterine ligaments. Jour. Amer. Med. Assoc., vol. 61, No. 14, Oct. 4, p. 1247. (17) Moritz, Manrrep 1914 The distribution and significance of the Para- metrium. Jour. Obstet. and Gyn., Oct.—Dec., vol. 26, p. 178. 1913 On the nature of the so-called ligaments of Mackenrodt. Journ. Obstet. and Gyn. of the British Empire, March, p. 135. (18) FotrHereitt, W. E. anp Moritz, M. 1913 The supports of the uterus. The Reviewer, British Medical Journal, Feb. 22. (19) CunnincHam, D. J. 1909 Textbook of Anatomy, 3d Ed. (20) Morris, H. anp McMurricu, J. PLayrarrR 1907 Human Anatomy. (21) Prersot, Gro. 1907 Human Anatomy. (22) Gray, Henry 1908 Anatomy, Descriptive and Surgical. 42 FRANK E. BLAISDELL, SR. (23) Emmet, Tuoos. Appis 1884 Principles and Practice of Gynecology, 3d Ed. (24) Scoauta, Friepricn 1896 Lehrbuch der gesammten Gynaekologie, Leipzig und Wien. (25) Somers, Gro. B. 1912 Utero-vaginal prolapse in elderly woman. Journ. Amer. Med. Assoc., vol. 58, No. 25, June 22, p. 1981. (26) Kerra, ArtHUR 1902 and 1913 Human Embryology and Morphology. (27) ParaMorE, R. H. 1910 Lancet, May, p. 1893. | | , | { | SPOLIA ANATOMICA, ADDENDA II ARTHUR WILLIAM MEYER From the Division of Anatomy of the Stanford Medical School TWENTY-TWO FIGURES CONTENTS A congenital intra-cranial, intra-dural adrenal..................2-.-0-eee02: 43 The relation of skeletal to body weight in the adult guinea pig.............. 50 Lymphoid nodules in the liver of Aluco pratincola...................-...... 51 TE OOPS TU SPPIPIGRE, <2. ook 750. Fig. 5 Relation of large intra-capsular chromaffine body to the adrenal. Until the direct connection with and indeed the inclusion in the reflexion of the capsula propria, of this muscle mass had been conclusively established by following the serial sections I was inclined to regard it as an accidental inclusion in spite of the suggestive staining reactions of the strand of connective tissue with which the portions first seen were so intimately associated. These muscle fibers are so well-preserved that the transverse striations are very plainly visible in some of them. Their loca- tion is indicated at ¢ in figure 2. Strands of the spinal portions =i SPOLIA ANATOMICA 49 of the accessory nerve are also contened in the capsule which also contains very wide exceedinsiy thin-walled vessels and a good deal of fat. It is very vell-preserved except in certain small areas. } he structure of this specinen and of the surrounding tissues make it very evident tha’!t 1s not metastatic in origin. Not the least indication of ma8nancy can be seen anywhere and the only possible explarUlon of its presence here, it seems to me, is that a small mas°f early embryonic mesnechyme was in some way included in ?¢ dura. This mesenchyme must then have differentiated ir? 2ll the constituents of the adrenal, into chromaffine be-®S; striated muscle, connective tissue and fat, the presence ‘ which tissues prompts one to put the specimen among terat@#ta. Various ~!ters on pathological anatomy seem to be agreed Maat the called primary hypernephromata occur only in the abdomin: ©2Vity. It is said that they are found but rarely in the "15; that they occur mainly in the kidneys but rarely main © liver, ovary, testes, ureter and the broad ligaments of the rus. Upon the advent of malignancy these aberrant adren’ My, to be sure, metastasize as any other malignant row ' any part of the body. None of the writers consulted haycclerred to or have themselves described anything at all rable to what is here reported, however, and were it not for © fact that this supernumerary adrenal is accompanied ‘iped muscle one might, in spite of the chronological diffi- ub even, be inclined to consider the possibility of later vortation or even of the migration of specific cell masses i! then formed the various complexes. But its location on polnal portion of the accessory nerve between arachnoid and dv together with the peculiar vascular relations which it yrubtedly must have had with the meningeal vessels, alone 2.) such a supposition quite untenable. Such a supposition wit also imply the transportation of all these various cell isl at least in part by the cerebro-spinal fluid, to this liar location. p E ANATOMICAL RECORD, VOL. 12, No. 1 50 ARCHUR WILLIAM MEYER The very small amount of medulla within the specimen with restriction of the sympathetic elements almost wholly to the aberrant and extra and intre-capsular isolated chromaffine bodies is remarkable however, espvcially in that location. From the appearance of the specimen one gets the impression that although the sympathetic elements attempted to penetrate the cortical mass they only succeeded in reaching the capsule and the surrounding connective tissue leaving the corpus proprium overwhelmingly epithelioid. THE RELATION OF SKELETAL TO BODY WEIGHT IN THE ADULT GUINEA PIG From computations based on statistics given by Donaldson (15) in table 53 the weight of the dried skeleton forns 1.94 per cent of the body weight in the new born, 4.38 per cent in the half grown and 4.09 per cent in the adult rat. Waldeyer (’10) gave the weight of the skeleton of two women of 40 years as 3.306 kg. and 3.585 kg. and that of a centenarian of 102 years as 1.185 kg. Although the body weights are not givn he weights given for these three fat-free skeletons indicate; that they formed from 3 to 6 per cent of the normal averagebody weight. From the accompanying table it is seen that the scele- ton of the adult guinea pig forms a somewhat smaller percenage of the body weight, than that of man but somewhat less tian that of the rat. One would I think, expect his from a mre comparison of the body forms of the rat and guinea pig. Since only ten guinea pigs were used for this determinaton the percentages obtained can, to be sure, not be regarded as being so near the actual for the guinea pig, as are those of Da- aldson for the rat. Nevertheless with three exceptions, the pe- centages obtained agree very well indeed, thus reducing tle probable error. Number 26 was not pregnant and hence lés fat. Pig No. 35 was in the early and pig No. 20 in the lae stages of pregnancy. Hence the divergences noted in thee cases may probably be accounted for by these things. Similer differences are also found in Donaldson’s large series and coulc, to be sure, be accounted for very easily by varying conditions 0 nutrition alone. ~ SPOLIA ANATOMICA ial All animals in this series of ten pigs were pregnant, save No. 26. The weight of the uterine contents was, however, always subtracted from the total body weight before percentages were calculated. The skeletons were cleaned by heating the fresh carcass in a 1 per cent solution of gold dust for five to seven hours. They were then dried for one week in a thermostat at a temperature of 54 to 55°C. after which the first weighing was done. Next they were placed in benzine for six to seven days in order to ex- tract the fat and dried at room temperature for a week. The second weighing was then done. The treatment was exactly the same in several groups which were handled together. From the percentages representing the relative weight before and after treatment with benzine it is seen that the reduction in weight amounts to 4 to 5 per cent of the weight of the cleaned, air-dried skeleton. This is a remarkably low amount of fat when compared with the results given for other animals. j DRY | | SKELETON DE- . = | pee. aes BODY WEIGHT SKELETON | PERCENTAGE /PRIV ED OF FAT PERCENTAGE 20 64 751.30 30.400 4.04 29).555 3.93 2 51 722.00 | 25.550 3.53 24.420 | 3.38 22 48 813.50 29.700 | 3.65 | 28.400 | 3.49 23 44 844.20 98.520 | .3.37 28.097 3.32 26 0 751.00 33.300 4.43 32.870 4.39 27 33 943.85 31.520 3.34 30.737 3.25 28 31 866.56 30.780 3.55 29.172 3.36 29 29 692.66 24.730° 3.57 | 93.490 3.36 31 25 901.50 30.790 3.41 29.875 | 3.31 35 15 688.30 28.840 4.19 27.770 4.03 LYMPHOID NODULES IN THE LIVER OF ALUCO PRATINCOLA In a specimen of the common barn owl—Aluco pratincola— lymphoid nodules of varying though small, size were found dis- tributed at random throughout portions of the liver. The lat- ter which looked wholly normal had been removed from a young owl about six months old. It contained no signs of inflamma- tion or degeneration, either macro- or microscopically, and the young owl which had been under observation for some months had never shown any signs of illness. 52 ARTHUR WILLIAM MEYER These accumulations of lymphocytes mixed with erythrocytes were of microscopic size, the largest measuring only a small frac- tion of a millimeter. Most of them were irregular in form and included extensions of the parenchyma of the liver into them Not rarely, however, one of the smaller follicles was surrounded by a very thin but distinct layer of connective tissue which could be regarded as a capsule. Those observed contained neither germinal centers nor an evident reticulum. The lymphocytes were crowded together and included rela- tively few erythrocytes. When small collections of erythrocytes were present they were usually segregated fairly well from the lymphocytes. The nuclei of the latter were not pyknotic but vesicular, as is the case within the lymph follicles of lymph nodes. No polymorphonuclear leticocytes or giant cells were seen in these follicles and no evidence of any phagocytosis was observed. The lymphocytes were not arranged in cords but were scattered about miscellaneously and no lymph vessels or sinuses were observed. Comparatively large blood vessels were, however, not infrequently seen near these nodules, penetrating them, and forming sinusoids within them. The general appear- ance is indicated in figure 6. LYMPHOID NODES IN STRIGIDAE From an examination of 31 species including 14 families Jolly (10) concluded that, among birds, lymph nodes are found only in some members of the group ‘Lamellirostres’ of the family Anatidae. Jolly failed to find them in Branta bernicla L. for example. Thirteen species of ‘Lamellirostres,’ all of which fall in the family Anatidae and one in the Phoenicopteridae were examined by Jolly. Ten in the former and one in the latter, viz., Phoenicopterus roseus Pall, contained lymph nodes. Of the other families examined none were found to possess lymph nodes. ‘These families included: Alcidae, 1 specimen; Colym- bidae, 2; Laridae, 1; Ardeidae, 2; Otididae, 1; Tetraonidae, 4; Columbidae, 1; Picidae, 1; Corvidae, 1; Alaudidae, 1; Sturnidae, 1; Fringilidae, 1; Strigidae, 1; Polyborniae, 1. Oe SPOLIA ANATOMICA aa The inclusion of a specimen of Strix flammea L. in this list attracted my attention because I had incidentally in the course of other investigations, come upon what I took for lymph nodes, in a specimen of Aluco pratincola (Strix pratincola) the common barn owl. These nodes were found in the abdominal and tho- racic cavities and were taken for lymph nodes when removed. Unfortunately, however, since they were incidentally removed and since two other owls were then available, the exact loca- Fig. 6 Lymphoid nodule from the liver of Aluco pratincola. The surround- ing hepatic parenchyma is merely indicated. 475. tion of these nodes was not specially noted at the time of re- moval. The specimens were fixed in Zenker’s solution, how- ever, and were found to be unmistakably lymphoid in structure. These nodes were cylindrical in form, 2 by 1 mm. in size and pale grey in color. This color may be accounted for by the fact stated by Jolly, that the sinuses of the lymph nodes of birds seldom contain enough blood to make the nodes look pink. Under low magnification no germinal centers or lymph sinuses were evident, although the parenchyma was an open one. In spite of the pale grey color of the gross specimens and the ab- 54 ARTHUR WILLIAM MEYER sence of blood-filled sinuses the stained mounted sections seen under low magnification suggested hemal nodes. Upon eloser examination it was seen however that this was due to the pres- ence of an unusually large number of large pink hyaline looking cells (fig. 7) which suggested erythrophages. These cells were scattered throughout the section of the node and contained vesicular nuclei which were generally located near the periphery. Although not contained in sinuses they are apparently somewhat comparable to those described by Jolly in the sinuses of lymph lig. 7 Portion of a lymphoid nodule showing distribution of polykaryocytes and large acidophile cells. X 750. a, capillary. nodes of birds, as containing cellular débris within their proto- plasm. The débris according to Jolly was composed of rem- nants of nuclei, erythrocytes and blood pigment all of which were ‘‘transformed into globular masses taking an acidophile stain.”’ Rarely polykaryocytes of the above type were also seen. These were extremely large and the protoplasm not infrequently contained what seemed to be remnants of nuclei. Sometimes larger, irregular masses which apparently had been formed by the coalescence of polykaryocytes were also found. These -— SPOLIA ANATOMICA 00 masses were far larger than any I had ever encountered before in any lymphatic tissues examined (fig. 8). Since distinctly or indistinctly outlined erythrocytes were, however, never seen within these cells one is lead to doubt whether these large cells were really erythrophages for the nuclei of the erythrocytes un- less destroyed before ingestion or extremely rapidly after that, should have been visible in some cells at least. Fig. 8. Portion of alymphoid nodule. X 515. a, capillary. CAPILLARY CAPSULES IN THE SPLEEN OF ALUCO PRATINCOLA During the examination of sections of the spleen of the owl, under low magnification, small groups and isolated syncytial- like masses with a small central opening with a circular or more or less elongated form especially attracted attention. These capsules characteristic of birds gave the impression that the spleen was studded with extremely large multinucleated giant cells, were always contained in the areas of lymphocytes and were absent in those areas of the spleen which were largely or almost wholly composed of erythrocytes. Upon higher magnification it was found that the small central openings con- tained in these masses were fine vascular capillaries which were 56 ARTHUR WILLIAM MEYER surrounded by a capsule of an epithelioid syncytium which was from four to six times as thick as the caliber of the capillary. Whenever some of these capsules were cut more or less obliquely and also when adjacent capsules coalesced, large irregular masses with smooth outlines resulted, but whenever the capsules were cut transversely the small capillary was usually centrally located in the section. Not rarely two small capillaries one of Fig.9 A group of capillary capsules of the spleen of Aluco pratineola. X 750. which was eccentrically placed were contained in such a cross . section. Some of these capsules lay directly beneath the splenic capsule or even caused it to bulge. They were always well- defined 2nd sometimes surrounded the capillary at the point of branching. Because of this fact and because of the consequent fusion of such adjacent and other capsules and perhaps also for other reasons, the size of these capsules varied considerably al- though there was but little variation in the ealiber of the en- . closed capillaries (figs. 9 and 10). > ~ SPOLIA ANATOMICA 57 Not infrequently two fairly concentric circles of nuclei were evident in cross section of these capsules. The cuter circle lay directly beneath the periphery and the inner directly around the capillary. The latter was, of course, formed by the nuclei of the endothelial cells of the capillary and except for a slightly smaller size and a somewhat more oval form the nuclei forming the inner and outer circles seemed to be identical in appearance. They were always vesicular and contained a number of dis- tinct chromatin granules. Sometimes they were distributed irregularly throughout the cross-section of the capsules. The inter-nuclear protoplasm stained pink with eosin and was non- granular. Cell boundaries were never recognizable and no cells except rarely a few isolated erythrocytes or fragments of such were seen in the syncytium of the capsules. In some cases the capsules instead of being composed of an epithelioid syncytium were composed of such merely along a narrow margin of their periphery and in the region immediately surrounding the capillary. The intervening space is partly filled with a rarefied tissue the individual nuclei of which are surrounded by small more or less confluent, amounts of proto- plasm giving these portions the appearance of mesenchyme. Aside from these capsules the entire absence of Malpighian corpuscles attracted attention. Not a single corpuscle was found in the sections examined and the only substitute for them were these large circum-capillarial capsules surrounded by lymphocytes. Since as many as a dozen of these capsules often lay quite closely together forming rather large pink-staining masses which were surrounded by blue-staining lymphocytes they were very conspicuous. The portions of the spleen exam- ined contained few large sinuses, a considerable quantity of erythrocytes, few trabeculae and had a thin capsule. Kyber (’70) found the capillary capsules in the dog to be 0.05 mm. wide and 0.15 mm. long. According to Kyber, Fenenko also described capillary capsules first so-named by Schweigger- Seidel, (62) who worked on the pig. They were discovered by Billroth, (’57) in birds and described by Miiller (’65) in frogs, 5S ARTHUR WILLIAM MEYER reptiles and birds. Bannwart (’93) found them in the cat and Kultschitzky (’95) in Putorius vulgaris. Kyber thinks that they are formed by local distensions (Auf- treibungen) of the adventitia of the arteries, which then enclose the splenic parenchyma in the form of a thin sheath of the ends of the terminal arteries. Kyber states that previously to his publication they had been described in the pig, dog, cat and hedgehog only, but Bannwart states that Muller found them indicated in the mole and rabbit also and found them in cap- sules but non-striated muscle and polymorphonuclear leucocytes were never noticed. The very large size of the capsules in the owl’s spleen as well as the large caliber of the capillary are evi- dent by a mere reference to the figures. PHAGOCYTOSIS IN THE LIVER OF FELIS DOMESTICA The animal from which this specimen was obtained was an old but well-nourished pregnant female. The four foetuses and one abnormal ovum were about 3 em. long. Although the cat had been handled very carefully and was killed in a gas chamber the abdomen was found full of fresh blood. Upon inspection of the viscera it was found that blood was oozing from the whole of the ventral surface of the liver and upon gently wiping the surface with a wad of cotton it was evident that the oozing came from small dark discrete points, the central veins. A little firmer wiping abraded the thin capsule and exposed the paren- chyma of the liver. The latter was very friable and yellow but the other viscera appeared macroscopically normal except that the ovaries were cystic. Upon microscopic examination the capillaries of the liver which contained but little blood, were found to contain numer- ous erythrophages as shown in figure 11. These cells which had a distinct cell membrane and a flattened crescentic nucleus which had been pressed against the cell wall, were about the size of the cells of the hepatic parenchyma. They were en- gorged with erythrocytes the outlines of which were still plainly visible in many of them. No phagocytosis was present in the spleen, sections of which showed a rather rarefied parenchyma. SPOLIA ANATOMICA 59 Sof Fig. 10 Two adjacent partly fused capsules containing a branching capil- lary. X 720. Fig. 11 Phagocytosis in the liver of a cat. & 1050. The kidneys showed a few old lesions but nothing else of conse- quence. In spite of the very active phagocytosis of erythrocytes in the capillaries of the liver of this cat I found no indication what- ever of phagocytic activity on the part of the hepatic parenchyma itself as observed by Browiez (’99). Nor did I see instances of 60 ARTHUR WILLIAM MEYER phagocytic activity on the part of the attached endothelial cells such as was observed by Heinz (’01). Nevertheless, the character of the phagocytic cells suggests an endothelial origin and to that extent confirms Kupffers (’99) conception of the phagocytic capacities of the endothelium of the liver. The entire absence of phagocytosis in the spleen of this cat would also seem to preclude an extra-hepatie origin of the erythro- phages in this case. Hemorrhage into the abdominal cavity and apparently always from the liver, was also observed not infrequently in other ani- mals killed by the use of illuminating gas. Since the animals are placed in a roomy lethal chamber and the gas turned on so slowly that death almost invariably results without a_ struggle, I can only suggest that for some reason unknown to me, extreme congestion of the liver with a possible change in permeability of the capillary and capsular walls must occur during death by the use of illuminating gas. THE ARCHITECTURE OF THE PROXIMAL EXTREMITY OF THE HUMERUS While scrutinizing the nature and the extent of the epiphyseal line in mature bones my attention was attracted to small areas adjacent to the epiphyseal line, in which the spongiosa is not infrequently absent. Often when not completely absent it is rarefied. These areas which were observed in the humerus, recalled Wards triangle and the similar rarefied areas in the spongiosa of the bodies of the vertebra and of the os calcis. Closer examination of a series of humeri showed that rarefaction or absence of the spongiosa were correlated with the retention of the epiphyseal line or plate. These absorption areas were always located in the lateral region of the shaft directly under the greater tuberosity a region in which the epiphyseal plate is best preserved. It is interesting and significant that a similar al- though not a corresponding absorption of the spongiosa can also be rarely seen near the epiphyseal line of the great tochanter of the femur. SPOLIA ANATOMICA 61 As shown in figure 12 these absorption areas occur on both sides of the epiphyseal plate and their size and the completeness of the absorption of the spongiosa, seem to vary directly with the completeness and strength of the epiphyseal plates. Whenever a part or the whole of the epiphyseal line was marked by a partial or complete bony septum or by two parallel thinner septa, the areas devoid of spongiosa were found the larg- est. Sometimes, however, there was only a partial absence or a rarefaction of the spongiosa and as shown in figure 12 when no epiphyseal line was evident there was no indication of absorp- tion. If on the other hand the epiphyseal line was absent al- together no absorption areas were found. ‘This relationship would seem to suggest that a strong epiphyseal plate relieves the spongiosa about it of most even if not of all, of the strains and stresses and thus causes its atrophy and rarefaction and finally its complete absorption. This conclusion would seem to be supported by the occurrence of all manner of transitions between a perfectly normal spongiosa and complete absorption and it is significant that the trabeculae of the spongiosa which are still preserved extend mainly at right angles to the epiphyseal plates thus acting as braces to relieve lateral strains. That the presence of such remnants of the epiphyseal plates has resulted in the absorption of the spongiosa is also suggested by the specimen of the humerus shown in figure 13. This spec- imen shows a large absorption area in the spongiosa, directly beneath the site of the great tuberosity, in which the spongiosa has completely disappeared probably in consequence of the disuse following articular disease. This humerus came from an extremity in which the long head of the biceps and the com- pacta in the region of the tuberosities had been completely destroyed by arthritis. There is no evidence of pathological processes within the spongiosa, however, Although the com- pacta of the humeral head shows some erosion it is also very evidently atrophic. Since the spongiosa around the empty area shows nothing suggestive of a pathological process it seems not unlikely that in this case absorption of the spongiosa may at least have been hastened by, even if not directly caused by, a lessened use during disease. * * . Ler) ee ye ait Pas ~~ 4 Fig. 12 Absorption near epiphyseal plates in three humeri and a great tro- chanter of the femur. 62 SPOLIA ANATOMICA 63 SERIAL TRANSVERSE BONY MEDULLARY SEPTA OF THE TIBIA von Recklinhausen (’93) described transverse partitions as not infrequently occurring in the broad and thick portions of the diaphysis of long bones where the compacta is thin. He states that as many as twelve such partitions may be present in the lower,end of the femur when the latter seems inflated (aufgetrieben) when viewed from the outside. According to von Recklinghausen transverse septa may also occur anywhere in the tibia and rarely also in the fibula and ulna, but never in the Fig. 13 Absorption area under the greater tuberosity in a case of chronic articular disease. humerus and in the upper femoral regions. They were also found regularly in the bones mentioned when multiple exos- toses were present, in femora with a short neck and, in general, in bones of light weight possessing the loose texture characteristic of osteomalacia. They were also found in bones from females below ten to twenty years and also in the upper extremity of the tibia of a twelve year old dog. von Recklinghausen emphasized that these septa may be distributed at regular intervals and may be found even in the middle of the shaft where the compacta is thickest. He concluded that they are remnants of the solid 64 ARTHUR WILLIAM MEYER portions of the bone which originally formed the epiphyseal plates of the growing bone, and have no mechanical significance. The occurrence of isolated complete, partial or fenestrated transverse septa in the shafts of certain long bones, is of course very common, but the presence of a series of parallel septa at comparatively short intervals as shown in figure 14 is rare. Fig. 14 Tibia with transverse septa. This specimen also deserves special comment because of the rarefaction and the nature of the spongiosa between the septa. The atrophy of the spongiosa near and between the septa can, it seems to me, be attributed to the presence of the trans- verse septa. Although none of these five partitions are com- plete and although all of them are fenestrated, rarefaction of the spongiosa is especially evident near them. All of these septa are found in the dorsal portion of the proximal extremity SPOLIA ANATOMICA 65 of the tibia and three of the five are much stronger than the other two which are composed merely of a framework of spon- giosa. The spongiosa between the septa is represented by a few very fine strands only and these extend mainly in a ventral direction at right angles to the septa. Nothing observed in this specimen militates against von Recklinghausen’s belief that such septa are remnants of former epiphyseal plates, but transverse septa located at or very near the midpoint of the shaft of a long bone could hardly be regarded as having such an origin. Moreover, the rarefied spongiosa in the proximity of the septa and also between them, indicates quite clearly that such septa are not necessarily or even very prob- ably, wholly without mechanical significance as von Reckling- hausen suggested. INIAL FOSSAE AND CANALS The skull shown in figure 15 I owe to the generosity of one of our former students Mr. Benjamin R. Hewitt. It was taken from an Indian Mound near San Jose, California, and as shown is markedly deformed. Although the deformation is marked it is nevertheless quite symmetrical and shows itself mainly by a decided flattening in the occipital region and of the vertex. The forehead is an extremely receding one, a gentle depression marks the glabella and the supra-supraciliary regions. The skull is decidedly prognathous and the norma frontalis which forms an angle of approximately 40 degrees with the vertical, is roughly parallel to the occiput. The obelion is located about 3 em. posterior to the line passing through the mastoid processes and is marked by two small pits about 3 em. apart which lie on opposite sides of the sagittal suture and which probably represent obliterated emissary foramina. The cerebellar fossae are deep, that on the right being the deeper, as usual. The floor of the left fossa is exceedingly thin and defective, partly no doubt from post mortem decay. The portion of the occipital bone bounding this fossa is much thinner, however, being only a few millimeters thick. All the sutures are still evident on the ex- terior and a good-sized ‘os Ineae’ is present. The linea nucha THE ANATOMICAL RECORD, VOL. 12, No. 1 66 ARTHUR WILLIAM MEYER Fig. 15 Indian skull with inal eanal a, rear; b, side view suprema is very evident but the linea nucha superior can not be dentified definitely. A very marked external occipital pro- tuberance is present. This protuberance takes the form of a SPOLIA ANATOMICA 67 torus 4.5 em. long and somewhat over 1 cm. high at its mid- point. One centimeter above the protuberance there is found a very marked but definitely circumscribed oval pit 1.2 by 0.7 em. deep. The canal leading from this pit is obstructed in part, by a plate of bone about 1 mm. thick the edge of which has very probably been destroyed. On sagittal section of the skull it is seen that the canal is somewhat irregular in form being obstructed by the thin plate of bone mentioned above, on the inferior portion near its inner orifice. The latter is irregular in form and measures 7 by 7.5 mm. Since the canal is funnel shaped the outer orifice is much larger, measuring 1.8 by 0.8 cm. in the transverse and vertical diameters respectively. The mastoid foramina are small, the parietal are obliterated but the jugular foramina are large. Hence it seems to me that one could hardly assume the presence of obstruction to the venous return and regard this canal as an enlarged occipital emissary vein. Moreover, were it to be regarded as such it would for several reasons be an extremely rare instance. The occipital emissary vein is usually small, it not infrequently pierces only one table and is often absent altogether. It is true that the canal in this skull leads partly into the stlctis for the left lateral sinus but the character of the canal itself is wholly different from the enlarged mastoid canals not infre- quently seen especially in rachitic skulls as emphasized by Merkel. It is possible, to be sure, that the canal in this skull has nothing in common as to its origin, with the sulci and fossae found in this region in the other skulls. Nevertheless if it is to be regarded as an enlarged emissary canal its character can only be explained by assuming a decidedly deforming influence upon it by the forces which deformed the skull. It is impossible to find a satisfactory embryological explana- tion for this peculiarity and since three other Indian skulls in a small collection of 60 possess, roughly similar depressions in exactly the same location, it is probable that these pits or de- fects have another origin. It is true that these defects lie in the region of the union of the interparietals with the supraoccipitals 68 ARTHUR WILLIAM MEYER but their character does not suggest a developmental origin. In one of the other three specimens there is a small circular depres- sion about 3 mm. deep at the center and 13 cm. wide. The other two skulls merely have very irregular small depressions and a fourth shown in figure 16 has a definite larger depression di- rectly in front of the superior nuchal line.! Pe, ee : wt “= Fig. 16 Indian skull with peculiar depression It is true that Frasseto (02) in a purely theoretical and hypo- thetical discussion says that an inial fontanelle was described by Maggi (00) and by Statirenghi (99). I have examined a number of papers by these authors but have been unable to find anything in their comparative anatomical studies at all com- parable to what is here described and pictured. ‘In view of Dr. Hrdliéka’s large acquaintance with skeletal remains gathered in widely different parts of the world and especially with American Indian re- mains, | brought the accompanying illustration to his attention. My expecta- tions were fully justified, for Dr. Hrdliéka has a series of specimens with similar, even if not identical, characteristics in the Smithsonian Collection. SPOLIA ANATOMICA 69 NOTES ON ‘SENILE’ ATROPHY OF THE CALVARILUM The erratic nature of bone resorption on the calvarium in the region of the parietals and sutures must continue to impress everyone. This is particularly true since we are in the habit of attributing the differences in relative degrees of senile atrophy between the bones of the upper and lower extremities, to dif- ferences in activity. Waldeyer (’10) also had recourse to such an explanation in connection with the findings in his unique study of the skeleton of a centenarian but in the case of the peculiar concentric atrophy of the calvarium we are left without this explanation. Voigtel (1804), Lobstein (’24), Rokitansky (’44), Virchow (54) and Maier (’54) were among the earliest investigators who described examples of this peculiar form of atrophy. Voigtel who refers to several earlier authors pictured a specimen from Meckels collections in which the atrophic area measured 3 by 2 inches. In one of the two cases reported by Virchow the bone in the atrophic area which measured 2.5 by 1.5 inches was, only “1 9 line thick.” According to Virchow the atrophy in the regions of the tubera parietalia never extends beyond the ‘linea semicir- cularis’ the insertion of the temporal muscle always definitely limiting the area. Virchow found atrophy present in other regions of the calvarium, however, and also noted joint changes. Maier who likewise described two specimens of calvaria reported a case of death following fracture in one individual. Maier like Virchow, emphasized the porosity of the whole cal- varium, the whiteness of the atrophic and the yellow color of the preserved areas, and spoke of the presence of a peculiar reticulated appearance due to the presence of lighter stripes among the yellow. Maier, however, found that the atrophic area extended beyond the ‘linae semicircularis.’ In the first skull reported by Maier the bone in the region of the ‘tuber parietalia’ was translucent over an area as large as a ‘Zwolf- kreutzerstiick;’ that is, about 3.5 em.; and as thin as ‘Post- papier.’ In the second skull the atrophic area was two inches long and one inch wide. 70 ARTHUR WILLIAM MEYER In reporting the case of a woman of 90 years who had suffered a fracture of the calvarium indirectly as a result of such atrophy, Humphrys (’90) also stated that ‘“‘The most common parts for the extreme thinning are the parietal bones on the side of the sagittal suture, midway between it and the tubera, causing the remarkable symmetrical depressions of which many specimens exist.’’ Humphrys also was impressed by these “‘changes of an opposite nature’ in old age—the absorption from without and the deposit from within. Rokitansky suggested a probable relation to lues but Virchow, Maier, Humphyrs, Ziegler, Aschoff and others all refer the atrophy to senility alone. Smith (06) called attention to the fact that this form of atrophy is rare in European crania and stated that Humphrys found only six instances of it in Euro- pean museums. According to Smith this peculiar form of atrophy is common in ancient Egyptians and never affects the parts of the calvartum covered by muscles. Smith further stated that a ring of bone 1 em. in diameter is always left around the parietal foramina. Although out of the 70 specimens exam- ined by him, not one was found below the age of 25 or 30 years, Smith nevertheless concluded that ‘“‘It cannot be regarded as a senile change because it frequently occurs in crania where the coronal, sagittal and lambdoid sutures show no trace of clos- ing.’ Although this atrophy was not found limited by sex, Smith found it present only in skulls taken from the tombs of the wealthy from the period between the fourth and nineteenth dynasties. Smith came to the conclusion that this atrophy is not congenital but is due to a continuous, slight pressure be- cause he found, ‘‘ This cranial thinning only in those people who were accustomed to wear wigs of enormous proportions and of great weight.’”’ Smith added by the way of qualification, how- ever, that a causal relation does not necessarily exist between the two. The first calvarium upon which I wish to comment is one with a roughly rectangular depression 3 by 5 em. long and 2 mm. deep over the mid-frontal region. The borders of this depres- sion are very regular and smooth and a roughly corresponding SPOLIA ANATOMICA ne bulging of the inner table is present but the two do not coincide exactly and cannot definitely be attributed to fracture. The portions of the coronal and practically all of the sagittal and lambdoid sutures which show on this calvarium are obliterated internally but are still evident externally. The thickness of the calvarium varies from 4 to 9 mm. and measures 5 to 7 mm. over the depressed area. The sulci of the middle meningeal artery are not deeper than usual but the arachnoidal and lacunar depressions are unusually large and deep, some of them extending well through the outer table. Yet on the whole this calvarium is heavy and its general appear- ance does not suggest senility. Compared with the measure- ments of Anderson (’00) which it is unfortunately very difficult to utilize because they are recorded in sixty-fourths (!) of an inch, this calvarium is above the average weight. The diploe are quite well-preserved but the lamina are thick. Only the right parietal foramen is preserved. The second specimen which is very evidently senile has a small absorption area over the sagittal suture about 2 to 3 em. anterior to the obelion, and a similar though less pronounced area on the lateral mid-parietal regions directly medial to the temporal ridges. The coronal and lambdoid sutures show faintly on the exterior and the whole anterior vault of the skull up to the absorption area in the mid-line shows definite vascular and nerve markings. The sulci for the right supraorbital nerve extend beyond the coronal suture. The internal surface of this calvarium is rough and shows considerable deposit. There are several exostoses in the frontal region and also deep tortuous arterial sulci. The condition of the rest of the skeleton would suggest that a marked reaction probably of syphilitic origin, was present. Although the frontal sinuses are small this calvarium measures over 1 em. in thick- ness. The lamina externa and interna are very thin but the diploe thick and well preserved. The coronal and lambdoid sutures are faintly indicated externally and the right parietal feramen is well-preserved. 72 ARTHUR WILLIAM MEYER The third specimen is a very light calvarium in which the location of the sutures is marked by sulci. There is a very shal- low and narrow suleus over the coronal suture but the sulci over the dorsal half of the sagittal and the lambdoid sutures are deep and wide and by their nature remind one at once of the peculiar large absorption areas in the parietal bones above re- ferred to. As shown very imperfectly by the photograph in figure 17 there are roughly-corresponding, comparatively large absorption areas in the posterior mid-parietal regions. Both Fig. 17 Calvarium showing absorption areas £ ~ these parietal absorption areas and the central sagittal area con- tain small areas varying from 1 to 4 sq. em. in which the bone is less than 0.5 mm. thick. This calvarium which is light and thin, measures 7 mm. in thickness at the lateral border of the parietal absorption areas. This is the greatest thickness found. xcept for a slight roughening along the superior sagittal suleus there is no evidence of bone deposition on its anterior. The figure formed by the sagittal and lambdoid absorption areas although T-shaped, has nothing in common with the T-sears of the dolmen skulls reported by Manouvrier (04). Since the latter have a mechanical origin being according to Obermeier, SPOLIA ANATOMICA 73 due to scraping for aesthetic or dedicatory purposes they have a wholly different character. The slightly pitted outer surface of this calvarium also suggests senility. The sutures and pari- etal foramina were totally obliterated. The fourth and most interesting specimen is from the body of a female 72 years old. The whole cadaver except the lower por- tion of the face, suggested senility and considerable loss of Fig. 18 Deep ‘senile’ excavations in the parietals with the accompanying brain. weight relatively shortly before death. The deep parietal ex- cavations shown in figure 18 attracted attention as soon as the body was unwrapped and recalled the descriptions of Humphry, Maier and Smith and an illustration in Ziegler (’02). Com- paratively slight sagittal pressure on either side in these areas, produced the peculiar crackling sound of thin bone. The scalp was not abnormally adherent anywhere but the calvarium was apparently very thin except perhaps along the 74 ARTHUR WILLIAM MEYER ridges which surrounded the depressions. The large frontal sinuses were outlined indistinctly through the thin lamina ex- terna. Upon removing the calvarium the dura was found de- cidedly adherent to the calvarium as is customary in old skulls. It was especially adherent over part of an area 2 by 1 cm. where the inner table of the right frontal sus was completely absent. The dura here fused with the sinal lining which was not espe- cially adherent to the bony wall. There was no evidence on this calvarium of bone formation, except in the region opposite the left pterion where a small rough hemispherical nodule 7.5 mm. at the base and 3.5 mm. high projected into the cranial cavity. This nodule which was imbedded in the posterior end of the mid-frontal gyrus, was somewhat adherent with its base, to an oval depression to a rough plate of bone about 6 by 10 mm. in size on the inner table. The rough scale-like character of the inner table in the region of the frontal sinuses, a few scat- tered much smaller plaques of bone on the inner table and the deep arterial sulci with steep walls all suggested the deposition of bone on the interior. As indicated by the depth of the fossae the absorption of the parietals is almost complete on both sides, their thinnest por- tions measuring less than 0.5 mm. over several small areas and less than 1 mm. over the whole of the rest of the floor of the depression which covered oval areas about 4 by 3em. These areas as measured on the level of the outer table were approxi- mately 8 by 5 em. large and were surrounded by a slanting wall about 1 em. high. Although the thickness of these surrounding bounding walls on both sides, measured from 9 to 12 mm.. the rest of the calvarium in the non-depressed areas was only 7 to 10 mm. thick. In addition to the marked absorption in the areas mentioned, that in the temporal regions was also very noticeable, yet this calvarium still measured 1.6 mm. in thick- ness, at a point 2.5 em. above the internal occipital protuber- ance. About 3 em. above the external auditory meatus it measured 3.8 mm. Although the thickness of the calvarium 1.7 cm. above the orbits was 1.7 em., the inner table which was SPOLIA ANATOMICA 5 absent in some places opposite the frontal sinuses was only 0.75 mm. thick and the outer only 1.25 mm. Numerous small areas of total absorption were also present over comparatively large areas of the tegmen tympani. The three largest of these areas in the tegmen measured 3 by 2 mm. and while those on the right side were smaller the tegmen never- theless looked honey-combed. Small absorption areas extend anteriorly quite close to the sulcus for the middle meningeal artery. The digital impressions and juga cerebralia were not very evident. The brain showed no very marked atrophy but the arachnoid was very thick especially over the whole frontal region. It was web-like and not unlike loose cotton in gross structure, about 4 mm. thick and very adherent throughout. After fixation in formaline the brain weighed 1035 gm. The stature of the body was 158.3 cm. The evidences of senility were not confined to the skull, how- ever, for the ribs were represented by a mere shell of very thin, pliable bone which could easily be compressed between the fingers. The costal cartilages except the first, however, showed no signs of calcification even in their interior. They contained only a few small grayish dots which had not, however, proceeded to the stage of calcification. The laryngeal and nasal car- tilages were not calcified either and the mandible was not markedly senile. The preservation of the latter was due to the fact that portions of the lower incisors, the canines, the left and right lower premolars, the roots of the first right lower molar and the stumps of two roots of the second right lower molar were preserved. That the roots of the absent teeth had not been lost very long before death is shown by the irregular- ities of the gums due to unabsorbed remnants of the alveolar processes. The rest of the skeleton is markedly senile. I have also been repeatedly impressed by the character of a general absorption occasionally seen in the posterior region of the parietals. Not rarely this absorption stops almost com- pletely when the lambdoid suture is reached thus causing the squama of the occipital to rise like a wall along the suture. It 76 ARTHUR WILLIAM MEYER is exceedingly difficult to understand what can be responsible for the character of the absorption in this region of the parietals. The lambdoid suture appeared normal in all these cases and we cannot have recourse to muscle action as preservative agents. There was no evidence of rickets in these cases. Although the senile character of such atrophies of the cal- varium as here reported has been and may be called in ques- tion, yet in these cases the atrophy probably was not due to the continuous pressure of a lesser weight, as Smith concluded for Egyptian skulls. Although the ages vary considerably the his- tories of these cases unfortunately are not available. The - fourth case was that of a woman born in this country. Hence weight bearing with the head, can be quite confidently excluded. Moreover, the erratic location of these atrophies as well as their peculiar shape makes it exceedingly difficult to conceive how pressure could be exerted upon those areas unless the head were born well-flexed upon the chest during exertion which would interfere seriously with respiration. Besides weight-bearing or pressure of considerable magnitude and for long periods of time do not result in such atrophy. THE RELATION OF BRAIN ATROPHY TO CRANIAL ATROPHY A case of very marked brain atrophy in an old woman, accom- panied by congenital porencephally might be expected to furnish some evidence even if slight, of the relation of cerebral atrophy to the thickness of the overlying calvarium. Since the de- crease in the volume of the cerebral cortex and the accompany- ing enlargement of the lateral ventricles do not cause a decrease in intra-cranial pressure it might be assumed that no deposit of bone should occur on the internal surface of the calvarium in consequence of brain recession. This is especially true since in cases of hydrocephalus an increase in intracranial pressure is accompanied by thinning of the calvarium and because con- stant pressure from other causes has a similar effect. Never- theless, thickening on the interior of the calvarium in the lateral frontal regions in consequence of deposit of bone from within as SPOLIA ANATOMICA 77 noted by Humphry, (’58) is very common in connection with atrophy of the frontal lobes in senility. But until more is known about the cause of deposition of bone on the inner surface of the ealvarium it would be wrong to assume that there is a causal relationship between the two. Especially since we do not know why in one case of atrophy there is marked absorption within with consequent thinning of the calvarium and decided broad- ening of the arterial sulci with their final obliteration through absorption, while in the other, on the contrary, there is just as marked a thickening in consequence of deposition within accom- panied by deepening of the arterial sulci to such an extent that large portions of the vessels may be almost enclosed. Fig. 19 Primary congenital porencephaly The brain showed in figure 19 was taken from the body of a woman 75 years old, dead of myocarditis. Neither the skull as a whole nor the calvarium showed any markedly senile changes. After fixation in formaline the brain weighed 1034 grams. How- ever, there had been some post mortem drying and shrinkage. The meninges were normal but all the cerebral arteries were somewhat sclerotic. Atrophy of the left hemisphere seems clearly more marked than that of the right. This is evident even along the longitudinal fissure, the frontal lobe and the lat- eral and central sulci. Marked asymmetry in the shape and the arrangement of the gyri also exists. This is especially notice- 78 ARTHUR WILLIAM MEYER able in the porencephalic area which lies in the posterior ex- tremity of the medial frontal gyrus and affects the latter and also the pre-central gyrus in the region around the posterior extremity of the inferior frontal suleus. The defective area measures 2.9 by 2.1 cm. and is approximately 2 cm. deep. The nature of the surrounding gyri as well as the condition of the corresponding area on the right side and that of the brain as a whole seem to indicate that this defect is not due to senile atrophy but to faulty development. This assumption was confirmed by section of the brain. There were no evidences whatever of lesions and the gray substance was as thick at the bottom of the porencephalic area as elsewhere. It was merely a case of failure of this area to develop properly and the defect is hence truly congenital. The peculiar arrangement and form of other gyri also suggests this. The arachnoid bridged over this depressed area and the overlying calvarium showed a gently rounded eminence on the interior and also a corresponding depression on the exterior. The calvarium here was also slightly thicker than the corre- sponding area on the other side but because of the correspond- ing depression of the outer table this increase in thickness was only very slight. The respective thicknesses at corre- sponding points on the two sides of the calvarium were 8 and 6 mm. The fact that the outer table was depressed over this area also indicates, it seems to me, that the local cerebral deficiency was 2 congenital rather than an acquired defect for, with the latter, one might expect a thickening of the calvarium from within unaccompanied by a depression from without. Although the left hemisphere plainly looked more atrophic than the right it nevertheless weighed 11 grams more. The measurements for length, breadth and width of the two hemi- spheres were practically equal, but considerable differences be- tween the volume of the two ventricles was found to exist. The volume of the right ventricle was 24 cc. and that of the left 20 cee. which accounts in part for the deceptive appearances. The volume of these ventricles is also considerably above the average found by Harvey (’09) who gave the ventricular volume for SPOLIA ANATOMICA 79 brains with an average weight of 1314 grams as 15.08 ce. for the left and 13.49 ce. for the right side. Considerable differences in ventricular volume were encountered, however, by Harvey. The character of this calvarium does itself not suggest senility. The parietal foramina are obliterated but the lambdoid suture is still preserved within, and the coronal sagittal and lambdoid sutures, especially the last two, are still well-marked externally. The whole inner surface of the calvarium is slightly rough, how- ever, plaques of new bone are found here and there and the region of the superior sagittal sinus is marked by a broad ridge which was apparently moulded upon the broadened longitudinal fis- sure. No absorption areas are present externally and the ar- terial sulci are not especially marked. Indeed, those over the depressed portion of the calvarium which covered the poren- cephalic area are especially shallow thus also suggesting that there has been no special deposit of bone there. The thickness of the calvarium in the lateral frontal region is 1.1 em. The diploe are sclerotic and the calvarium which has a thickness of 5 mm. in only a few places, is heavy and strong. The clinical history of this case could not be obtained but a careful examination of the entire body revealed no evidence of defective muscular development or of asymmetrical atrophy as a result of developmental conditions or of paralysis. Sec- tion of the cord showed it to be symmetrically formed and no gross signs of degeneration could be detected. The greatly emaciated condition of the cadaver was rather puzzling and remained unexplained until the stomach was opened. It was somewhat dilated but otherwise normal save for the presence of a papilloma directly in front of the pyloric orifice. This tumor which was 3 cm. long arose from an en- larged base which lay partly upon the gastric margin of the pyloric sphincter but which was not adherent to the gastric musculature. The body was 1 em. wide and 0.5 em. thick being flattened dorso-ventrally and the free portion distinctly papil- lamatous. The pyloric musculature was not hypertrophied and the base of the tumor was alone sufficient to completely obstruct the ori- SO ARTHUR WILLIAM MEYER fice. Since the papilloma was freely movable its inclination toward the antrum probably did not prevent it from being forced against the pylorus and thus further obstructing the passage of gastric contents into the duodenum. Hence it seems highly probable that this woman suffered much from gastric trouble and that her emaciation was largely due to the mechanical diffi- culty caused by the tumor. THE CAUSE OF SOME LARGE PARIETAL EMINENCES It is usually stated that large pariteal eminences are due to a local displacement outward of both laminae of the skull. In the great majority of cases this explanation holds but in case of Fig. 20 Outline of a cross section of calvarium with a large parietal eminence the calvarium outlined in figure 20 the large parietal eminence on the right plainly has a different origin. As the figure shows, the inner table of this skull has suffered no outward displace- ment whatever and the prominent eminence is due to a consid- erable thickening and a gradual deflection of the outer table. Sclerosis of the diplce is present and several areas of dense con- nective tissue and what looked like red marrow, are interpolated between the two tables the space between which is largely filled in by compact bone. This calvarium which was taken from the body of a man 70 years old at time of death, is heavy partly beeause sclerosis is so complete in the frontal region and also because the calvarium SPOLIA ANATOMICA 81 as a whole shows only slight evidence of absorption. Although the arterial sulci are only slightly deeper than normal, very evident thickening in the interior in the lateral frontal region is nevertheless present. The greatest thickness of the right parietal eminence is 15.5 mm. and the thickness of the corresponding point in the left is 8.5 mm. The greatest thickness in the left lateral frontal re- gion is 1.0 cm. and 0.9 cm. on the right. The thinnest point in this calvarium is in the right temporal region where the section measures only 2.5 mm. yet in spite of the thinning in this region the whole calvarium does not even remotely suggest senility. It is true that all the sutures are well obliterated and that the sagittal suture is marked by a very slight suleus which extends over the metopic suture which can be recognized still. These things are, however, of but little moment. The right parietal foramen is still evident and there is no cae of concentric atrophy in that region. The periosteum was not especially adherent over this bosse which measured 6 by 4 em. in the segittal and parietal planes respectively. Its surface is very smooth and it looks denser; that is whiter; than the rest of the calvarium but there is no good evidence of the presence of a pathological process and the rest of the skeleton was wholly normal. EXTENDED FUSION OF THE SECOND AND THIRD RIBS Although more or less extensive bifurcation of ribs is rela- tively common fusion is much rarer. In fact Bland-Sutton (’99) stated that fusion of the ribs resulting in the formation of bi- cipital ribs occurs only in connection with the cervical ribs or ‘in relation to the first and second ribs. JDisse (’96) made a similar statement. Dwight (’07) however, held that a bicipital rib may occur also by the fusion of the first thoracie with the second beyond the tubercles. Merkel (99) in a fine summary, merely states that anomalies occur at the posterior ends in the region of the tubercles where the ribs may send processes toward each other and a similar statement is made by Nicolas (’11) THE ANATOMICAL RECORD, VOL. 12, No.1 82 ARTHUR WILLIAM MEYER and Thompson (714). In Rauber-Kopsch (’11) it is stated that now and then it is observed that the margins of two or more ribs may fuse for a greater or lesser distance. This statement was also made by Thompson (713) who merely said that fusion of adjacent ribs may occur. In the second edition Thomson re- ferred to Meckel. Campbell (’69) reported a remarkable case of union of a num- ber of ribs by cartilage and also by bone accompanied, however, by multiple exostoses and also by the formation of bone in the diaphragm and of cartilage in the right lung. The presence of so many exostoses and the occurrence of bone formation in the soft tissues makes it probable ,however, that, in this case, the union of the ribs was due to a pathological rather than a de- + velopmental process. Turner (’70—’71) referred to a case of fusion of the first and second ribs reported by Hunauld in 1740. Hunauld is also said to have possessed a fetal skeleton from the seventh month in which the upper five left ribs were united posteriorly and in . which the sixth and seventh ribs were also partially united. In this article Turner pictured two specimens of fused ribs which from comparative anatomical considerations, he apparently de- clared to be fused cervical and first ribs. In a later article Turner (’82—’83) concluded that the two specimens in question were fused first and second ribs. A fuller account than that contained in any of the above texts and handbooks is that given by Lane (’83). Lane gave several instances of fusion of a cervical with a first rib and of fusion of the first and second ribs. He found the common shaft formed by the union of the first and second ribs in one of his cases, 14 inches broad dorsally with a maximum width of 24 inches. The latter point was located 11 inches behind the termination of the lower rib. Unfortunately Lane did not give the length of the union but judging from his drawing this was as long as the breadth of the united ribs or 2} inches. Although the ribs in this speci- men were said to be firmly united the ‘‘outer surface of the shafts still remained very prominent after fusion, being separated from one another by a deep groove. The inner surface of the com- SPOLIA ANATOMICA 83 mon shaft was smooth, however, and presented no irregularity or ridge of any sort.’”’ Lane remarked that this specimen is peculiar in its great breadth and in the thinness and incurvation of its lower part. Two similar cases were previously described by Turner, and Bryce (’15) stated that Valenti (03) also described a case of fusion of the second and third vertebra accompanied by ‘appar- ent’ fusion of the second and third ribs for the greater part of their length. The most comprehensive study of the occurrence of variations in ribs is that by Hrdlicka (00) who examined over 1600 ribs Fig. 21 Fused second and third ribs and numerous Indian skeletons. In an examination of this large amount of material; Hrdlicka found only one ease of junction of the third and fourth ribs but three cases of the much more com- mon anomaly—tusion of the first and second ribs. The specimen shown in figure 21 was taken from the cadaver of an old man, is composed of the second and third ribs. The rest of the skeleton was normal and the thorax was not unus- ually asymmetrical. Hence the anomaly was not very evident and my attention was called to it by Messrs. Supple and Tufts, two of our students. These two ribs which are of normal length were fused throughout three-fourths of their length. Neverthe- less their relations were maintained exactly for they had inde- pendent cartilages and a short bifurcated medial extremity. 84 ARTHUR WILLIAM MEYER Unlike the specimens of fused first and second ribs described by Lane, the individual shafts are almost completely obliterated in the medial half of the fused portion, both within and without. This is true of a rhomboidal area lying within 8 cm. of the sternal extremity of the second and 7 cm. of the same extremity of the third rib. The intercostal groove is only very slightly preserved on the external surface of the lateral half of the fused portion. Since the fused portion extends to within 9 mm. of the medial extremities the appearance here is that of a bifurcated rib. The common shaft is 12.3 em. long, the first rib 22.8 em. and the second 26.5 em. The width of the fused portion is 3.5 em. laterally, and 4.3 em. medially but the distance from the superior border of the first rib to the inferior border of the second rib at the medial extremities is 6.1 cm. This increase in width is due to the fact that the short non-fused medial extremities diverged rather markedly to meet the costal cartilages. The distance from the mid-point of the head of one rib to the midpoint of the head of the other is 2.1 em. or normal. In the lateral area of fusion the second rib is very definitely outlined for a distance of 4.3 em. and the third for a distance 7.9 cm. Throughout this region of only partial obliteration of the shafts the intercostal region is marked by a broad deep sulcus and the outlines of the individual shafts are well preserved ex- ternally but not internally, where the intercostal region is marked by a decided ridge which gradually disappears toward the medial extremity. This rounded ridge looks not unlike the inner sur- face of a rib and gradually merges with the shaft of the first rib. The subcostal suleus of the second rib is fairly well marked to a point of 2.5 em. beyond where the fusion begins but that on the first rib extends 4.5 em. beyond this point. The greater length of the sulcus on the first rib is due to the fact that it lies external to the bony union between the ribs for a distance of 4.3 em. The intercostal nerves ran internal to the fused portion. ‘The superior margin of the flat, fused area is much thicker than the inferior, the two measuring 6 and 2.5 mm. respectively. The greater thickness of the superior margin is due to the better development of the bony area which would be comprised by the SPOLIA ANATOMICA 85 second rib. In thickness, spacing and form where separate, and also in length where fused the ribs are entirely normal. The intercostal muscles are of course absent in most of the fused area which is covered by a strong intercostalmembrane. The muscles are present, however, in the lateral portion of the fused area where the external portion of the bodies of the individual ribs are only partly unaffected by the fusion.’ BILATERAL ABSENCE OF THE EXTENSORES CARPIULNARES Such a variation as the above is not mentioned by Le Double (97) or in the larger handbooks such as those of Bardeleben and Poirier et Charpey. Neither do Quain nor any of the other German and English textbooks consulted speak of it. Gruber (85) and Turner (’85), however, each reported a somewhat sim- ilar case. Gruber also says that he could not find anything like it in the literature. Gruber observed the specimen reported by him in 1883, in the left arm of a young male 20 years old. It was the only specimen observed by Gruber in the course of the personal dis- sections of 600 arms! In this case the muscle was represented merely by a tendinous strand which extended, however, all the way from the external condyle to the fifth metacarsal. This strand which was fused with the fascia was 5 mm. wide above, 3 mm. wide and 1 mm. thick in the middle of the forearm and 4mm. wide and 2 mm. thick at the point of insertion. The lower portion was supplied with a vaginal sheath and lay in an ulnar suleus of normal dimensions. Other fairly common mus- cular variations were also present in this arm. Turner who referred to Gruber said the case observed by him was wholly comparable to that of the former. In Turner’s case also, the sulcus on the ulna was normal although the ten- 2? Through the courtesy of Dr. Hrdlicka, it has since been my privilege to examine specimens of fused ribs in the Smithsonian Collection, and if 1 remem- ber correctly, one of the specimens is very similar to what is here described. Through the courtesy of Dr. Lamb I was also enabled to examine a specimen of extended fused ribs in the museum of the surgeon-general, which belongs still lower down in the series of ribs. 86 ARTHUR WILLIAM MEYER dinous slip was only one-sixth the size of the normal tendon. Turner emphasized that such an anomaly was not mentioned by Macalister or Testut in their works on muscular anomalies, and that this case was the only one observed by him in thirty years’ experience in the dissecting room. A very careful examination of both upper extremities of this — male subject by Messrs. Supple and Tufts who noted the ab- sence early in their dissection, and by Professor Congdon and myself did not reveal a trace of these muscles. It seemed to me at first that a remnant of the tendon had fused with the internal lateral ligament of the wrist but a comparison showed that this ligament varies sufficiently in strength and distinctness to justify one in including those on these arms among the normal variations. Although the extensores carpi ulnares muscles were completely absent in these arms the sulci on the dorsal surface of the distal extremities of the ulnae, in which they lie were nevertheless present and practically normal in size and depth. Hence these specimens again illustrate a certain independence in the forma- tion of these and similar sulci, which nevertheless may be moulded by tendon pressure even if not primarily due to them No other anaomalies were found in this cadaver and no unusual strands or thickenings were present in the fasciae of these fore- arms. No modifications could be determined in the fifth metacarpals. UNILATERAL ABSENCE OF THE TWELTFH RIB IN AN ORANG- OU-TANG In a skeleton of an adult orang-ou-tang which seems to possess no other unusual characters the condition of the ribs deserves a word of comment. There are eleven pairs of. ribs which look normal except for the development of a flat wedge-shaped square bony process one centimeter square on the anterior upper sur- face of the tenth rib. This process which is 3 mm. thick at the base arises from the superior costal border a few centimeters ventrally from the angle and must have come close to the inner surface of the superior rib although the latter bears no sign of such contact. Many of the ribs on both sides also bear tri- angular bony extensions along their inferior borders in the re- SPOLIA ANATOMICA 87 gion of the angles a condition which may, however, for all I could learn, not be uncommon in the orang-ou-tang. The right transverse process of the nineteenth, or first lumbar, vertebra looked, practically like the corresponding process in human skeletons. The accessory and transverse processes were very well differentiated and were marked by a deep sulcus. The mammillary process though slender was well-developed and was separated by a broad deep sulcus from the inferior processes. The left transverse process, on the other hand, had the typical form except that its extremity was pitted by a very definite articular cavity which received the head of the twelfth rib. The latter was 8.8 cm. long and the eleventh ribs 18.2 em. It is particularly interesting that although the twelfth left rib did not articulate with the body of the vertebra but arose from the transverse process the right transverse process nevertheless showed no enlargement whatever and possessed a better dif- ferentiated accessory mammillary and transverse processes than most human vertebrae. THE EFFECT ON THORACIC FORM OF COMPLETE DESTRUCTION OF ONE LUNG Although the expression ‘‘One lung is gone”’ is not unfamiliar to medical students and to physicians who deal with large numbers of tubercular subjects it is necessary to recall that those words as customarily used by clinicians are not intended to be taken literally. Moreover, when one considers the great vessels in the root of the lung such a thing seems quite impossible. It would seem that death from hemorrhage must occur long before the great pulmonary vessels so near to the heart can be oblit- erated. Yet such a case recently came to my attention. The cadaver was that of a man beyond middle age and nothing which could be identified with the naked eye as pulmonary tissue remained of the left lung. The remnant of the pleurae was roughened and thickened considerably and the remnant of the root of the lung, was represented merely by a short stub of dense connective tissue containing the sclerotic and wholly, or partially obliterated extremities of the bronchi, blood vessels and calcified lymph nodes. The left thoracic cavity contained 88 ARTHUR WILLIAM MEYER but very little coagulum and was otherwise completely empty. The left dome of the diaphragm was but slightly displaced. The fact that the mediastini and the contained viscera were displaced so slightly is likely explained by the thickening of the left parietal pleura long before the lung was completely de- stroyed. I have not sufficient knowledge of either tuberculosis or pathology to venture a statement on the probable sequence of events in this case but the fact that the external form of the thorax was altered so slightly that it attracted no attention, does seem to indicate that not much tension could ever have been exerted on the thoracic wall as a result of an adhesive pleurisy. I recall that Hutchinson found that the long-expressed idea that tubercular individuals are flat-chested is erroneous but these findings do not imply that tuberculosis never affects thoracic form. Hutchinson found that the anterior posterior diameter in normal adult males between the ages of twenty to forty-four is 71 if the transverse diameter at the level of the nipple is taken as 100. This same diameter in 82 clinically tubercular subjects was found to be 79.5 and in 30 flat-chested individuals 80. Instances in which the thoracic form has been profoundly changed especially in cadavers in which the ribs are markedly senile, are, of course, frequently seen in dissecting rooms where most of the material used is from the senile or tubercular or from the senile and the tubercular. But it is difficult to see how a lung can be completely destroyed by tuberculosis without an attendant pleurisy and consequent long-continued tension on the chest wall as a result of fibrosis. Nevertheless, the conditions were as here represented and there can be no doubt about the tuber- cular nature of the disease in this particular case. OSSIFLICATLON IN THE ARACHNOID The occurrence of small hardened areas in the spinal arachnoid is comparatively common in dissecting room cadavers. Be- cause of this fact and also because these areas are usually very small I have in the past usually taken them for calcifications. SPOLIA ANATOMICA | 89 This assumption was based not only on their appearance but also on their physical properties. Some months since my atten- tion was called to certain small areas of apparent fibrosis of the arachnoid and to other very much larger horny plaques. Mr. H. M. Winans and Miss Dorothy Wood, two of our medical students noticed these peculiar plaques in the lower dorsal region upon exposing the spinal cord. The largest of them has an area of 2.8 by 1.6 cm. and the next largest 2 by 1.4 em. Both were less than 0.5 mm. thick, however, and were moulded so as to surround the dorsal portion of the cord in the region over which they lay. They were adherent neither to the pia nor dura. Both these large plaques and the smaller similar ones were very flexible and quite translucent. In cutting off a portion for microscopic examination it was evident that these plaques both had the hardness of bone rather than of horn and that they lacked the friableness or at least the brittleness of calcified areas. Examination of paraffine sections showed that these areas were composed of lamellae laid parallel to the flat surfaces of the plaques. The lamella contained some rather atypical bone corpuscles and were penetrated at intervals by perforating or atypical Haversian canals which here and there opened upon the surface. Several small narrow cavities were also found and the internal surface also showed a narrow calcified layer. The outer portion, on the contrary, was formed by a single layer of investing cells. The body was that of an Irish woman of 75 who had died of arterio-sclerosis and in whom the porencephalic brain referred to on page 76 was found. In view of the close relationship between fibrous tissue and osteogenetic processes, the mere presence of bone in the arachnoid is nothing particularly surprising. A discussion of this subject in man and animals with references to the literature and numer- ous interesting personal observations is given by Cushing and Weed (’15). 90 ARTHUR WILLIAM MEYER CUTANEOUS PIGMENT IN THE SPERMATIC FASCIA Although small quantities of cutaneous pigment are foundjin the cutis of man I am not aware that the so-called superficial fascia has been found pigmented. It is true that Toldt (713) found that the epidermis in the dark skin spots in Macacus inuus and Cebus libidinosus contained very little pigment and the deep layers of the corium much, but this pigment was intra and not intercellular for it was contained in large branched pigment cells. Toldt also emphasizes the relatively great pigmentation Fig. 22 Spermatic fascia from Didelphys virginiana in the corium of these mammals as compared to the lower verte- brates. Adachi (’03) on the other hand concluded that there is a correlation between the intensity of epidermal and dermal pigmentation but found that the quantities of pigment contained in the cutis were relatively small. Breul (’96) and Frederic (06) strangely enough found pigmented places in the corium most common in the relatively unpigmented regions. In figure 22 a portion of the spermatic fascia of an opossum is shown. The surface marked S is the superficial one and that marked D the deep one. As shown in this figure large masses of pigment are scattered about in almost the whole of this fascia SPOLIA ANATOMICA ; 91 the superficial layers of which contain thick layers of pigment. In addition to these larger masses pigment granules were also scattered about in the fascia but practically all of the pigment was extra-cellular. Although I am not here concerned with the finer questions relating to the origin and occurrence of normal skin pigment there can be little doubt that we are dealing with normal cutaneous pigment. The gross character of the over- lying skin alone makes this certain. The scrotum and the para-scrotal skin of this opossum was intensely bluish black. The color of this area made it look like ‘tache bleuatre,’ the peculiar shade being of course due to corium pigment. Since smilar pigmented areas as found in the scrotal region of this opossum are relatively common in mammals and since other similar areas were found on this very animal the scrotal pigmentation attracted no special attention until the skin and some of the superficial fascia had been reflected, butcher-wise, and the testes were seen to be just as intensely bluish black. On incising the spermatic fascia and reflecting the parietal tunica vaginals with it, the testes were seen to have the usual color. 92 ARTHUR WILLIAM MEYER LITERATURE CITED Apacut, B. 1903 Hautpigment beim Menschen und bei den Affen. Zeitschrift fiir Morphologie und Anthropologie, Band 6. von Ascuorr, L. 1911 Pathologische Anatomie. Zweiter Band, Specieller Teil, Jena. Anperson, R. J. 1900 Note on the comparative thickness of the skull as an index of brain recession. Internationale Monatschrift fiir Anatomie und Physiologie, Band 17. Browicz, T. 1899 Intussusception der Erythrocyten durch die Leberzelle und die daraus méglichen Bilder der Leberzelle. Anzeiger der Akademie der Wissenschaft, Krakau, 1899. 1898 Ueber intravasculire zellen in den Blut capillaren der Leber- acini. Anzeiger der Akademie der Wissenschaft, Krakau, and Bul- letine International de l’Academie des sciences de Cracovie, 1899. Bryce, Tuomas H. 1915 Osteology and arthrology. vol. 4, Part 1. Quain’s Elements of Anatomy, London. BreveEL 1896 Ueber die Verteilung des Hauptigments bei verschiedenen Menschenrassen. Morphologische Arbeiten, Band 4. BANNWART 1891 Untersuchungen iiber die Milz. 1. Die Milz der Katze. Archiv fiir mikroskopische Anatomie, Band 38. CaMPBELL, Joun A. 1869-70 Note of a case of abnormal union of several ribs. Journal of Anatomy and Physiology, vol. 4. CusHiInc, Harvey, AND WEED, Lewis H. 1915. Calcareous and osseous de- posits in the arachnoidea. John’s Hopkins Hospital Bulletin, vol. 26. Dwicut, THomas 1907 Human anatomy. Piersol. 1st edition. Philadel- phia, p. 153. Disse, J. 1896 Skelettlehre. Handbuch der Anatomie des Menschen. Bar- delben, Lieferung I. Donatpson, Henry H. 1915 The rat, data and reference tables. Philadel- phia. Frasseto 1902 Sur les fontanelles du crane chez l’homme les Primates et les Mammiferes en general. Essai d’une théorie topographique. An- throp., Paris, T. 13. Frepertc 1906 Zur Kenntnis der Havtfarbe der Neger. Zeitschrift fir Morphologie und Anthropologie, Band 9. GruBER, WenzeL 1885 Mangel des Musculus ulnaris. Archiv. fiir patho- logische Anatomie, Band 99. Harvey, Ricuarp 1911 The volume of the ventricles of the brain. Anat. Rec., vol. 5. Heinz, R. 1901 Ueber phagocytose der Lebergefiisse-Endothel. Archiv fiir Mikroskopische Anatomie, Band 58. Hropxricka, Ates 1900 Contribution to the osteology of ribs. Proc. Assn. Am. Anat., 14th Annual Session. Baltimore. Humpurys, G. M. 1889-90 Senile hypertrophy and senile atrophy of the skull. Jour. of Anatomy and Physiology, vol. 24. 1858 The human skeleton. London. 1889 Old age. Cambridge. : . | . SPOLIA ANATOMICA 93 Jouty, J. 1910 Recherches sur les ganglionslymphatiques des oiseaux. Archives d’Anatomie Microscopique, Tome 2. von Kuprrer, C. 1899 Ueber Sternzellen der Leber, Verhandlungen der anatomischen Gesellschaft. Jena, 1898, 12. Kyser, Epwarp 1870 Ueber die Milz des Menschen und einiger Saiigethiere. Archiv fiir Mikroskopische Anatomie, Band 6. Kuurscuirzky, N. 1895 Zur Frage iiber den Bau der Milz. Archiv fiir Mikro- skopische Anatomie, Band 46. Le Dovuste 1897 Traité des variations du system musculaire de l’homme. Paris. LogssTeEIN 1824 Traité d’anatomie pathologique. Tome 1. Lane, ARBUTHNOT 1883 Cervical and bicipital ribs in man. Guy’s Hospital Reports, vol. 42. Mater 1854 Beitrige zur pathologischen Anatomie. Eine Form der Schidel- atrophie. Archiv fiir pathologische Anatomie, Band 9. Manovvrier, M. L. 1904 Incisions, cauterisations et trepanations craniennes de l’epoque neolithique. Bull. et Mem. de la Soc. d’Anthrop. de Parise 5; L. -v. MERKEL, FriIepRICcH 1899 Topographische Anatomie. Braunschweig, Band 2, Badal Miuier, Wituetm 1865 Ueber den feineren Bau der Milz. Niconas, A. 1911 Traité d’anatomie humaine. Poirier P, et Charpey, P. Paris. Tome 1, p. 397. OBERMAIER, Huco 1912 Der Mensch der Vorzeit. Berlin. Ravusper-Korscw 1911 Lehrbuch der Anatomie des Menschen. 9. Auflage, Leipzig, Abtheilung 2, s. 44. von RECKLINGHAUSEN, F. 1893 Ueber normale und pathologische Architektur der Knochen. Devtsche medizinische Wochenschrift. Roxiransky 1844 Handbuch der pathologischen Anatomie, Band 2. Surron-Buanp, J. 1899 Human anatomy, Morris. Second edition. Phila- delphia. Smiru, G. E. 1906-07 The causation of the symmetrical thinning of the parie- tal bones in ancient Egyptians. Journal of Anatomy and Physiology. London. ScHWEIGGER-SEIDEL 1862 Untersuchungen iiber die Milz. Tuompson, Peter, 1914 Human anatomy, Morris. 5thedition. Philadelphia, p. 132. Tuomson, ArTHUR 1913 Human anatomy, Cunningham. 4th edition. New York, p. 277. Turner, Str Witttam 1882-83 Cervical ribs and the so-called bicipital ribs in man in relation to corresponding structures in Cetaceae. Journal of Anatomy and Physiology, vol. 17. 1884-85 Absence of extensor carpi ulnaris and presence of an acces- sory sural muscle. Journal of Anatomy and Physiology, vol. 19, p. 33444. : Toxtpt, K. 1913 Ueber Havtzeichnung bei dichtbehaarten Saugetieren ins- besondere bei Primaten, nebst Bermerkungen iiber die Oberflaichen- profiliering der Siugertierhaut. Zoologische Jahrbiicher, Band 35. 94 ARTHUR WILLIAM MEYER VaLENTI, G. 1903 (Title not ascertainable. Cited after Bryce.) Mem. d. Acad. d. Se. d. Instit. d Bologna, Ser. 5, T. 9. Vircuow, Rupotr 1854 Kleinere Mittheilungen, 111. Ueber die Involution- skrankheit (Malum senile) der platten Knochen. Verhandlungen der Wiirzburger physich-medizinischen Gesellschaft, Band 4. VorcteL 1804 Handbuch der pathologischen Anatomie, Band 1. WEIDENREICH, FrRANz 1910 Das Gefiisssystem der menschlichen Milz. Archiv fiir mikroskopischen Anatomie, Band 58. WaupeYer, WiLHetm 1910 Das Skelett einer Hundertjahrigen. Sitzungs- bericht der Preussischen Akademie der Wissenschaft. ZincLeR, Ernst 1902 Lehrbuch der speciellen pathologischen Anatomie. Zehnte Anflage, Jena. THE HYPOPHYSIS OF THE GUINEA PIG C. M. VANDERBURGH From the Department of Anatomy of Stanford University SEVEN FIGURES The guinea pig was used as the object of this study because it is readily obtainable and also because though thousands are used yearly for experimental purposes, this animal has been neglected from an anatomical standpoint. The only reference found to the anatomy of the hypophysis of the guinea pig, was one by Oppel (14) who referred in a few lines to its general relations and another by Paulesco (08) who quotes Fischera in regard to the weight of the gland. The general relations of the hypophysis of the guinea pig _ are very similar to those existing in other vertebrates and its microscopic structure is quite similar. The most striking dif- ference found was the presence of cilia in the cleft and also in the epithelial cysts or vesicles of the glandular portion. Ac- cording to Trautman (’09), ciliated epithelium has been reported in the vesicles in the hypophysis of man and the rabbit, but Trautman was unable to find any in the cysts of any of the domestic animals studied by him. These included the horse, ass, cow, calf, sheep, goat, hog, dog and cat, and although he could not be certain Trautman thought he saw ciliated epithe- lium in the lining of some portions of the cleft of the pituitary of the hog. In the guinea pig this ciliated epithelial lining of both the cleft and the vesicles was found uniformly present some- where in all the specimens examined, but it was not present in all cysts nor did it completely line the clefts. Within the latter, in fact, it did not appear at all on the side adjacent to the pars intermedia and only on portions of the opposite side. These portions are easily recognizable even under low magni- 95 96 . C. M. VANDERBURGH fication by the large, clear, columnar cells, which often project into the lumen, as little mounds. In some cases, however, the ciliated portion of the lining epithelium extends over con- siderable stretches. The fact that ciliated epithelium is pres- ent in both the cleft and in the closed vesicles of the epithelial portion seems to indicate that their origin is the same and that both undoubtedly are remnants of the lumen of the epithelial pouch (Rathke’s pouch) developed from the buccal epithelium. In several cases, for example, it was possible to trace the lumen of typical vesicals to the lumen of the cleft. METHODS In every case a portion of the brain was removed with the hypophysis in order to retain its relations, particularly with the third ventricle. All microscopic preparations were cut serially in paraffine. Sections cut sagitally in the cranio-caudal plane were used mostly since they best show the relations of the different parts. Zenker’s fixative, followed by methylene blue and aqueous eosin were found to be the best general fixa- tive and stain. Flemming’s and Orth’s fluids were also used. The phosphotungstie acid-hematoxylin stain was found valuable in staining neuroglia and ependymal fibers and cells, and also reticular connective tissue. Next to the methylene blue and eosin, aqueous eosin and iron-hematoxylin were found to be most satisfactory. The latter stains the cell boundaries es- pecially well (figs. 3 and 4). OBSERVATIONS The hypophysis of the guinea pig is comparatively large. Measurements made upon three freshly killed adult animals show that it weighs between 22 and 32 mgm., varying somewhat in different specimens. Paulesco (’08) quotes Fischera as giv- ing the weight of the gland as 0.015 gram, but he does not state whether his data were derived from specimens of fresh or preserved glands. By weighing the brain also in each case, HYPOPHYSIS OF THE GUINEA PIG 97 it was calculated that the gland represents between 0.52 per cent and 0.70 per cent of the total brain weight. The measurements made on formaline fixed glands as to size gave the following averages: mm. sETNISVErSely;|(WIGeSAPAlu) Sacer ssa. cess aiin tal cic ox0s acs ofaieis octet eae o 5 aie -CAUCHILY (WIGERE DALE) ac.cco cu Ie oe oo Fx clad aninite Cacia tvs vinta « 4.2 DWorBo-veneral live (Widest part) oc. < = nae \ = et T,n0s ~* ral SS on at me ie wo ee a = x< = uJ. = er -Dil x CLATTENBURG 13 134 49} THORACIC DUCT OF ADULT GUINEA PIG a a ee 5 + = 75 17 A. CLATTENBURG H. 136 CLATTENBURG H. A. 138 ey CS eS a a te = OP A Lin R 1. y ‘ wa —— a ak oe Q oS Ni OBSERVATIONS ON THE SWEAT GLANDS OF TROPICAL AND NORTHERN RACES ELBERT CLARK AND RUSHKIN H. LHAMON Department of Anatomy, University of Chicago TWO FIGURES The present preliminary report on the sweat glands was be- gun as a part of a joint investigation suggested by the late Paul C. Freer, Director of the Bureau of Science of Manila. The gen- eral problem was the supposed untoward effect of a tropical resi- dence on the white man and the supposed constitutional adapta- bility of the dark races to a tropical climate. The investigations were undertaken in a codperative way by chemists and physi- cists of the Bureau of Science, the departments ot Anatomy, Pharmacology, Physiology and Physics of the University of the Philippines and the United States Army Medical Board for the Study of Tropical Diseases. Certain claims of Daubler (’00) and Aron (’11) seem to indi- cate at least one definite adaptation of the dark races to the tropics and lead us to investigate the sweat glands. Daubler states that the size of the sweat glands of the native of tropical Africa is much greater than that of the European. Aron finds that the sweating apparatus of the aborigines of the Philippines, the Negritos, is much superior to that of the white man. This superiority he says is shown by the difference in the manner of sweating rather than in the amount of sweat produced. Accord- ing to Aron, the Negrito secretes small beads of sweat over the entire body, which soon forms a thin film. As the whole surface of the body is covered by this water film, the maximum cooling effect from evaporation is obtained. In the case of the white man, on the other hand, the sweating is practically limited to certain areas of the body surface. In these areas the sweating 139 140 ELBERT CLARK AND RUSKIN H. LHAMON may be quite profuse, but, as most of it drops off, comparatively little cooling effect from evaporation is produced. He suggests that the Negrito has a greater number of sweat glands which are more equally distributed over the entire body. We have attempted to compare the sweat glands of the tropi- cal races with those of the northern races. Our observations at present scarcely extend beyond a comparison of the number of sweat glands in certain definite skin areas of various races. Similar comparisons of other areas will be made as material is collected. A comparison as to size is an almost endless task as is shown by the work of Huber and Adamson (’03). These in- vestigators have found, from measurements of reconstruction a great variation in the size of the sweat glands in the Caucasian. The length of the tubule in the coiled portion of the gland from the plantar region of the foot of an adult was 4.25 mm. A gland from the hairy portion of the pubic region of a woman was found to measure 10.4 mm. They have further shown, and we have confirmed their findings, that it is almost impossible to de- termine with any degree of accuracy the size of sweat glands without making reconstructions of them. It is even now and then exceedingly difficult in a series of sections to trace with certainty a single gland, especially toward the beginning and end of a series of sections of any one gland. Loops of neighboring glands are often in close proximity and are apparently surrounded by a common layer of somewhat denser connective tissue so that a separation of tubules belonging to two, or now and then even three, contiguous glands can be made with certainty only by reconstruction. In maceration preparations of skin from the palmar region and from the chest in both American and Filipino we have noted a great variation in the size of the glands in each piece of skin taken. Frequently a given gland was fully twice as large as its neighbor. It is thus apparent that a racial comparison of sweat glands as to size to be of any pretence to accuracy must be based upon measurements of a vast number of glands in different portions of the skin of the several races. SWEAT GLANDS; TROPICAL AND NORTHERN RACES 141. TECHNIQUE In our study of the number and distribution of the sweat glands we find that there is less variation in those occurring on the plantar surface of the foot and the palmar surface of the hand than in other regions of the body. The sweat glands first make their appearance in the embryo in these regions. These areas also lend themselves readily to an extensive enumeration of the sweat glands. Our observations so far have been made principally on these regions. In a warm climate prints of the fingers, palms, toes and plantar surfaces of the feet can be made which in many cases will give a negative impression of every gland duct in the area printed. We have employed the method which is in use in the recruiting office of the United States Army. A very thin layer of the best printer’s ink is rolled out with a small hand roller upon a glass plate. The subject’s finger (hand or toe, etc.) is carefully and lightly pressed first upon the ink, and next upon a special type of glazed paper. The impressions. when made under suitable conditions, are remarkably clear and the duct of every sweat gland can usually be accounted for. Skin with much cornified epithelium will not make a satisfactory print unless pains are taken to macerate with a dilute caustic and scrape off the surface tissue. The gland ducts may be counted directly upon the hands and feet with the aid of a good hand lens of about 12 diameters mag- nification. This is facilitated by first rubbing a little powdered graphite upon the surface of the skin to be examined. This method is far more tedious and has been used only as a check upon the print method. The print method is obviously not adapted to those areas of skin where hair and wrinkles occur. As the ducts of sweat glands frequently open into hair follicles the glands can not be counted by direct inspection, Here we have resorted to maceration methods and to stained sections of skin cut parallel to the surface. The latter method was used by Krause in his study of the number of sweat glands in the skin of the various regions of the body. Fig. 1 Photograph of a finger print of the distal phalanx of the fourth finger (left) of an American white soldier. X< 6.5 SWEAT GLANDS, TROPICAL AND NORTHERN RACES 143 Fig. 2 Drawing of a finger print to show orifices of sweat gland ducts. X 10. Through the courtesy of the surgeon-gener: al’s office United States. Army and of the office of the ¢ chief surgeon, Philippien Division of the United States Army we have had the opportunity of examining finger prints of 300 American white soldiers, 200 144 ELBERT CLARK AND: RUSKIN H. LHAMON American negro soldiers, 150 Filipino (Christian) soldiers and 100 Moro scouts. In addition we have ourselves made many prints of fingers, toes, palms and plantar surfaces of the feet of Americans, Filipinos, Igarotes, Negritos and Hindoos. In counting sweat glands a glass slide with a graduated square (0.5 em. each way) was placed over the print, graduated surface down, and the specimen magnified 10 diameters. In good prints, as stated above, every sweat duct can be counted (fig. 1). The graduated square was always placed over the print of the distal phalanx in counting the glands of the fingers, and over that area where the cristae cutis form a whorl or delta. RACIAL VARIATION IN SWEAT GLANDS We have counted 248,998 sweat glands in } square centimeter areas of 1,572 fingers and 38,736 in the palms, toes and plantar surfaces of the feet. Table 1 shows the average counts per square centimeter of skin area for the various races. Rather uniform variations have been observed in the distribution of sweat glands in the fingers. The number of glands varies in dif- ferent areas of the volar surfaces of the fingers. The number is greatest near the distal ends and smallest in the immediate region of the flexion groove at the joints. As stated above, the TABLE 1 Average number of sweat glands per square centimeter of skin area in the various races as shown by fingers FINGERS OF LEFT HAND FINGERS OF RIGHT HAND 58 NATIONALITY j aes 5th | 4th 3rd_| Index| Index} 3rd 4th 5th Bz American (white)..........| 569.6'600.4)564.4/511.2 519.21546.4 590. 4/552. 4/558. 2 American (negro).......... 605. 2/634. 4/586. 0/576. 6)561. 6/590. 4/631. 2/594. 8597.2 EULLIDINIO p= siete oe aT 6 oe 652.0/691.2\653.6618.4.610.4/630. 4/692. 8/637 . 2/653 .6 VU ON Oto osp0 0 esto ee Ee 674.4)755.2|682. 8/650 .8)651 2/665. 2\725.6 64081684. 4 Negrito (adult)............| 670.4|752.0)718 .8/682. 4/666 . 0/720 . 0/736 .8|701 6/709. 2 Joho (0 ee Peaee yet ...| 722.0|813.6|744.0/657. 6/650 .0/737 .6|782 .0/738 . 4/738 .2 Negrito (youth).....<:..:.. 1010.0.996.0.942.8'942.0/865. 2/961. 2/982. 8/881. 2/950.0 Average per finger.......| 707.67|/749.9\696. 3/661 . 4/646. 2/693 .0)734.5'678.0 Gemeral averages ne.aa ss fein cise IGE, Sots E ok a1 0 ba ee Ee 698.5 SWEAT GLANDS, TROPICAL AND NORTHERN RACES 145 number is most nearly constant in the region of the whorl or delta than in other regions. Comparatively few prints of the thumb have been examined. In all of these, however, the number of sweat glands has been distinctly lower than in any of the other fingers. Of the other fingers the second or index finger has shown the lowest average number of glands per unit of skin area, while the fourth or ring finger has shown by far the great- est number of glands. A more detailed comparison of the num- ber of sweat glands per square centimeter of skin for the dif- ferent fingers in the several races is given below in table 1. The average number of glands per square centimeter of skin area for the finger for all the races examined is 624.4.! The greatest number was found in prints of the fingers of Negrito children. It was found that the number of glands per unit of skin area for the hands, feet and toes bears a rather close racial relation to the number on the fingers, and in the different individuals varies directly with the number on the fingers. As regards the differ- ent races our results show a greater number of sweat glands in all the tropical than in the northern races. Taking the American white soldier as the standard, the num- ber of sweat glands per unit of skin area was found to be 6.83 per cent greater in the American negro soldier, 16.61 per cent greater in the Filipino soldier, 22.34 per cent greater in the Moro soldier, 26.81 per cent greater in the adult Negritos, 31.72 per cent greater in the Hindu and 69.82 per cent greater in the Negrito youths and children. Additional details of these counts will be found in table 1. The greater number of sweat glands per unit area with the Negrito youth and child is no doubt due to a corresponding difference in size of the individuals. As all the sweat glands are fully formed at birth? it is merely a 1 This average is much lower than the estimation of earlier authors, thus— ‘Uber die Menge der Kniueldriisen haben wir iltere Angaben von Krause senior denen zufolge ihre Zahl zwischen 400-600 (Riicken, Wange, erste zwei Ab- schnitte der unteren Extremitaten) und 2600-2736 auf I 0” Haut schwankt und die grésseren Zahlen an der hanffliche und Fussohle sich finden. Neure Zihlungen von Hérschelmann ergeben viel naher stehende Grenzzahlen von 641 Fussrucken, und IIII (vola manus) auf I 0 em und viel mehr driisen.’’—Koelliker. 2 At the fourth month according to Wilder (716). THE ANATOMICAL RECORD, VOL. 12, No. 1 146 ELBERT CLARK AND RUSKIN H. LHAMON question of the increase in skin area during growth bringing about a dispersion of the glands. The ratio of the number of sweat glands in the American white soldier (100 per cent) to that of the Filipino soldier (116.93 per cent) and the Negrito adult (124.05 per cent) shows a wider variation than the dif- ferences in size* of the individuals of these respective groups. The American negro soldier is of the same approximate size as the American white soldier, and there are 6.83 per cent more sweat glands per unit of skin area in the former. The Hindus examined were of a larger average size* than the American soldiers and showed the highest sweat gland count for adults (131.76 per cent). Thus racial variation in size does not account for the difference in ratio of the sweat glands per unit of skin area. The number of sweat glands was determined in a similar manner from prints of the palmar surface of the hand and the plantar surface of the feet of Americans (white) and Filipinos. Successful prints were made from these areas of 6 American uni- versity men and 6 Filipino students; 325 separate areas were counted, giving a total of 38,736 glands. The average number of sweat glands on the palmar surface was 438.0 per square centi- meter in the American, and 473.6 in the Filipino. On the plantar surface of the feet there were 436.4 glands per square centimeter in the American and 498.4 in the Filipino. On the plantar surfaces of the toes there was an average of 527 and 525.5 sweat glands per square centimeter in the American and Filipino respectively. Thus the number of sweat glands in the Filipino was in this series 8.1 per cent greater on the palm and 14.1 per cent greater on the plantar surface of the feet than in corresponding areas in the American. In all these counts there was very little individual variation. Different indi- viduals of the same group gave almost the same count. * Captain Davis of the recruiting office of the United States Army in Manila tells us the average weight of the American white soldier is approximately 150 pounds and of the Filipino scout approximately 130 pounds. The Negritos are smaller and can be estimated at about 120 pounds. * All these Hindoos were tall, large and portly and averaged 160-165 pounds in weight. SWEAT GLANDS, TROPICAL AND NORTHERN RACES 147 We are not able to confirm Aron’s observation that the tropi- eal aborigines secrete only small beads of sweat over the entire body. On two tramping expeditions in the mountains of the Philippines which we were fortunate enough to arrange with a number of Negritos we observed streams of sweat running down the back, and copious sweating on scalp, forehead and face and sweat dripping from the chin. When making finger prints in camp it was necessary repeatedly to dry off droplets of sweat from the fingers of the Negritos. From the few maceration preparations mentioned above we are not able to discern any difference in the size of the sweat glands of the American and the Filipino. As regards number, al] our observations show a higher count in all the tropical races. These counts, furthermore, were made on those areas in which the number of sweat glands is the most nearly constant. BIBLIOGRAPHY Aron, Hans 1911 Investigation on the action of the tropical sun on man and animals. Phil. Jour. Se., Sec. B, vol. 6, p. 101. DdvBLER 1900 Die Grundziige der Tropenhygiene, Berlin, 1900. Cited by Aron. Huser, G. Cart and ADAMSON, Epwarp WILLIAM 1903 “J j , ' . " - . . 7 -, =. > . ‘ La « i q : ‘ » _ >, ) THE SIGNIFICANCE OF THE LUNULA OF THE NAIL MONTROSE T. BURROWS The Department of Pathology, The Johns Hopkins University, Baltimore, Maryland TWO FIGURES A structure, the significance of which has always been de- seribed by a series of different answers in modern text-books of anatomy is the naillunula. This is a small sharply circumscribed opaque area which is visible near the root of the nail of many individuals. Toldt assumed that the greater opacity of this region is due to the fact that the nail is covered here by a layer of matrix, the cells of which are undergoing active division. Unna expressed the opinion that the opacity is conditioned by the existence in this region of keratohyaline (omychogenic sub- stance). Ranvier has given a similar explanation (quoted from Poirier and Charpy, Traité d’anat. hum., Paris, 1901). He ascribed it to the existence of onychogenic granules which fill the cells of the rete mucosum. Y. Brunn, on the other hand, ascribed it to the fibrous structure of the cells undergoing keratinisation in this region of the nail. Although these particular explanations have for a long time appeared to be inadequate, it is of interest that no one, so far as the author has been able to determine, has attempted to study more carefully the peculiarities in the structure of this region. The fact that the lunula is marked off sharply from the remain- ing portion of the nail is a direct argument against several of the above explanations. Again, the large number of nails which one sees removed in the dispensary do not show that portion of the nail corresponding to the lunula more opaque than surrounding parts. During the last few months a careful study of the structure of the nail and its environment has been undertaken for the 161 THE ANATOMICAL RECORD, VOL. 12, No. 1 162 MONTROSE T. BURROWS purpose of establishing, if possible, some of those conditions which regulate its continuous growth throughout the life of the indi- vidual. This study has been planned to include not only a care- ful investigation of the morphology, histology, and the develop- ment of the nail but also a detailed study of its connective sup- port and vascular supply as far as they apply to the solution of the problem in question. It represents a continuation of those studies of the mechanism of cellular growth which have been carried on for some time in the laboratory by means of the tissue culture method. It was during the course of the early part of this study that the author noticed certain peculiarities in the structure of the nail in the region of the lunula which have appeared to be im- portant in conditioning its opacity. Since the lunula itself has a certain clinical as well as a morphological importance, and since these particular structural peculiarities have not hitherto been emphasized as important in the anatomy of any part of the body, although probably very significant as a part of the general mechanism of the growth of this structure, it became of interest to report them in a separate communication. The nails of the hands of five individuals have now been studied. They have included those of two adult white men, an adult negro woman and two white children. Lunulae were noted at the base of uncovered portion of the body of the nails of all the fingers of the white men and on the thumbs of the children. None was seen on any of the fingers of the negro woman be- fore the epitrichium had been removed. When the epitrichial layer is lifted, however, the sharply defined opaque lunular zone is seen. It is present in all the fingers of this individual. A number of these fingers were dissected at once. Others were fixed in formalin, dehydrated and decalcified. The blood vessels of a number of the fingers were injected with India ink. These fingers were likewise fixed in formalin, dehydrated and decalcified. The fingers which had been fixed were either em- bedded in colloidin, sectioned and stained or they were sectioned free hand, examined at once and later after they had been cleared. All the unfixed fingers were hemisected along the SIGNIFICANCE OF LUNULA OF THE NAIL 163 median sagittal plane. This was accomplished by first cutting through the nail and the soft parts with a sharp knife, the bone being later cut through with a fine saw. In every one of these fingers examined it has been noticed that the matrix of the nail underlying the lunula is not firmly adher- ent to the connective tissue stratum. Throughout the whole lunula area, the two layers do not apparently adhere but lie only in contact with each other. The slightest distortion of a sagittal section of the finger leads to the separation of these two layers. The open space thus formed is sharply circumscribed. It extends from the tip of the root of the nail, at which point the matrix is adherent, to a line which corresponds to the distal margin of the lunula zone. Beyond this line and over the whole body of the nail the matrix is firmly adherent to the underlying connective tissue layer (figs. 1 and 2). The connective tissue underneath the lunula is peculiar in structure. The fibrils near its outer edge run parallel to the lower margin of the nail matrix. They form a dense boundary layer or sheath. The connective tissue here is not vascular. The capillaries are reduced largely to a single or double layer which lies close to, and on the surface of this layer of connective tissue. The underlying portions of the connective tissue contains few capillaries. The matrix is also peculiar in this region. Its lower margin is in most parts straight. One sees very few indentations or papillary extensions into the underlying connective tissue. This is in sharp contrast to the outer pink portion of the nail body. Here the matrix is indented both longitudinally and _ trans- versely by connective tissue papillae. The connective tissue in this region is also strikingly different from that of the lunula. It contains many vertically placed fibrils which end directly at the edge of the epithelial cells of the matrix. They adhere closely to these cells or a basement membrane. The papillae as well as the neighboring portions of the connective tissue con- tain large numbers of capillaries. They form a dense network which fill the papillae and neighboring parts of the connective tissue. This dense capillary network is continuous with that 164 MONTROSE T. BURROWS of the lunula portion, the change from one form into the other taking place gradually under the distal margin of the lunula. A number of the nails of the fingers examined have been re- moved by stripping them back from their outer tip. By this procedure it is possible to remove the nail with the matrix of the i 2 Fig. 1 A thick sagittal section of the finger of awhiteman. A piece of paper has been passed through the opening between the matrix and the connective tissue. Fig. 2 A thick sagittal section of a finger of a white man which has been fixed, dehydrated and decalcified. The open space between the matrix and the connective tissue layer is readily discernible in the area corresponding to the lunula. lunula attached to it. Throughout the body of the nail the matrix is invariably torn. In none of the nails thus removed has the lunular portion been found to be more opaque than the remaining portions. Quite the reverse, this area is frequently more translucent. In a number of the nails which had been SIGNIFICANCE OF LUNULA OF THE NAIL 165 previously fixed, dehydrated and decalcified, the tip of the root does show a definite opacity. In none of these cases, however, is this opacity seen as far forward as the distal margin of the lunular area. It fades and gradually disappears a short distance distal to the tip of the root. Furthermore, this opacity is not noticed in all the fixed specimens. From these latter observations it seems evident that the opacity of the lunula is not conditioned by any peculiarity of the structure of the nail itself nor its matrix. Moreover, that the decrease in the capillary bed of the lunular area is not wholly, if at all, responsible for the opacity may be deduced from the fact that its change in density at the distal margin of the lunula is never as abrupt as the boundary line of the lunula demands. Further proof against this peculiarity playing any important role in conditioning the opacity is given by comparing the appear- ance of the lunular area with a portion of the body from which the blood has been removed by pressure. The lunula has a defi- nite opacity, while that portion of the body from which the blood has been removed has a greyish translucency. In the absence of other possibilities, the author has been led to believe, therefore, that the lunular opacity is the result of a reflection of the light at the surfaces of the junction of the matrix and the connective tissue of this portion of the nail. In the outer portion of the nail where the matrix adheres closely to the underlying connective tissue the light is transmitted di- rectly to the capillary bed giving it its characteristic pink color; while in the region of the lunula the well formed non-adherent surfaces of both the connective tissue and the matrix reflect the light. More direct proof of this assertion may be readily ob- tained by pulling a portion of the body of a living nail loose from the connective tissue, thus forming such a surface in this region. During the past few weeks the author has been performing work which has led to the production of this injury. In each instance when it has occurred the detached portion has shown anopacity quite indistinguishable from the opacity of the lunula of the same finger. It is of interest, however, that unless the injury is extensive the nail will again after a few hours become adherent and assume its former pink color. 166 MONTROSE T. BURROWS The decrease in the capillary bed of the lunular area has been described (Poirier and Charpy, Traité d’anat. hum., Paris, 1901). No one, on the other hand, as far as the author has been able to determine, has described the peculiarity of adhesion between the matrix and the connective tissue in this region, nor the relation of this peculiarity in structure to the lunular opacity. It will be of further interest to ascertain whether many of the patho- logical opacities of the nail are not the result of similar changes, and to study more carefully the conditions which lead to the ad- hesion of the matrix in outer portion as contrasted with that of the lunular zone, and the general changes in the connective tissue and in the arrangement and density of the capillaries in these two regions. The nail is one of those peculiar parts of the body which continues to grow throughout the whole life of the body. A more careful investigation of its general structure and the chemical and physical changes which accompany its growth may lead to many facts of importance for ultimately ascertaining those conditions which bring about and regulate the growth of other body structures. A PRELIMINARY EXPERIMENTAL STUDY ON THE RELATION BETWEEN MITOCHONDRIA AND DISCHARGE OF NERVOUS ACTIVITY B. TALBOT STRONGMAN Anatomical Laboratory, Johns Hopkins University We have heard a great deal about mitochondria in nerve cells, but unfortunately their study has not emerged from the purely descriptive into the experimental stage. The only record of experimental work on mitochondria in the nervous system is a brief note by Luna (’13, p. 415) on the changes in mito- chondria in nerve cells following section of their peripheral proc- esses. He found that the first changes consisted in a loss of the regular distribution of mitochondria, in an increase in their volume and in an increase in their affinity for iron hematoxylin; while in the more advanced stages of degeneration the mito- chondria disappeared completely. Archimede Busacca (’15, p. 232), working on the eye, found that mitochondria in the nerve cells of the retina increased in number on light stimula- tion. Accordingly it is a very pertinent question to inquire whether there are alterations in mitochondria of nerve cells in muscular fatigue. White mice were selected for the experiments because they are the smallest mammals which can be conveniently used in the laboratory and on account of the fact that more is known of the quantitative (Thurlow ’16) and qualitative (Nicholson ’16) variations in mitochondria in their nervous system than in that of any other animal. They were fatigued by the very simple method, which Prof. Tamao Saito uses, of letting them swim in water until they are exhausted. It was soon discovered that they swim better when the water is slightly agitated and is raised to body temperature. Otherwise they soon learn to float and refuse to exercise. 167 168 B. TALBOT STRONGMAN The experiments were controlled in the usual manner by using only mice of known age and by examining, in exactly the same way, an unexercised mouse of the same litter for comparison. Five experiments of this kind were made, the mice swimming from one to two hours continuously before they were completely exhausted. (Larger mammals will swim for a day or more be- fore exhaustion). Experience showed that young mice’ twenty- five to thirty days old are more suitable than adults because they are more easily tired. In each of the five experiments the fatigued mouse and the control mouse from the same litter were chloroformed. They were then fixed by the injection of a formalin and bichromate mixture in accordance with the method advised by Cowdry (’16, p. 34). The brains were removed, mordanted, washed, dehy- drated and cleared together in the same bottle. They were embedded in the same block of paraffin. Sections, cut 4 » in thickness from the fatigued and from the control, were mounted on the same slide so as to avoid variations in the staining which might otherwise occur. The sections were then stained with fuchsin and methyl green, the fuchsin coloring the mitochondria crimson and the methyl green staiming the Nissl substance a bluish green color. Preparations were made in this way of the cortex, the cerebellum and the spimal cord from each of the five experiments. The mitochondria show a slight numerical increase in the fa- tigued animals in three of the experiments, but the other two experiments do not show it, indeed, in one of them there is a definite decrease in the number of the mitochondria; in any vase, the variations in the number of mitochondria fall well within the range of variation which was found to be normal for the species. Neither could any definite change in the form of mitochondria be seen in the fatigued animals. True, in the Purkinje cells of the cerebellum some of the mitochondria were swollen up to form spherules (0.5 » to 1 uw in diameter), but, on careful ex- amination some of the controls showed examples of the same phenomenon, though in less marked degree. MITOCHONDRIA AND NERVOUS ACTIVITY 169 There was a tendency toward a clumping of the mitochondria in the cytoplasm of the fatigued animals as contrasted with the more or less uniform distribution in corresponding nerve cells of the control. This clumping, when it occurs, is particularly apparent in Purkinje cells at the base of the large dendrite in close relation with the ‘capuchon chromatique’ of Cajal. The Nissl substance is well fixed and is stained brightly with methyl green by the technique employed. A mild degree of chromatolysis was noticed in all the fatigued animals, in variable amount. It was usually most marked in the Purkinje cells of the cerebellum and in the pyramidal cells of the cortex. This would be inclined to lead one astray were it not for the fact that one of the normal control animals showed just as extensive chromatolysis if not more extensive than any of the fatigued ones. Vacuoles and clear, tortuous, unstained canals (the canalicu- lar apparatus of authors) were of common occurrence in both the fatigued and the control brains. In two of the experiments they were certainly wider and more clearly cut in the anterior horn cells of the fatigued than in those of the normal, but, again, this difference was not apparent in the other three. The nuclei and nucleoli were given careful attentionand proved entirely negative as far as a definite change is concerned in the fatigued animals. Unfortunately preparations were not made by the neurofibrillar methods and by the positive methods for the demonstration of the Golgi apparatus. Chromophile cells could be seen in both the fatigued and the control brains. They were never observed except in the higher centers, that is, the cerebral cortex and the cerebellum. The chromophile cells stained uniformly red with fuchsin (Cowdry 16, p. 36). If anything, they were slightly increased in number in the fatigued animals. It is evident, therefore, that the net result of these five ex- periments, with their controls, is to show that the mitochondria are surprisingly constant in nerve cells, rather more so even than the Nissl substance which is supposed to be concerned with the activity of nerve cells ‘per se.’ A fair degree of fa- 170 B. TALBOT STRONGMAN tigue, as well as a certain amount of fright, brmgs about no constant changes in them. It is certainly no longer necessary to take elaborate precautions in composing the minds of ani- mals before killing them for studies on the mitochondria in the central nervous system. It is quite possible that more pro- longed exhaustion will lead to definite and precise alterations in mitochondria in nerve cells. But it would appear that the study of mitochondria in fatigue in muscle cells, thrown into isometric tetanus outside the body, is more likely to bring about changes in them. Mitochondria occur in all nerve cells though we have very little idea of what their true significance is. Certain it is, how- ever, that they are stable structures which are not essentially altered with slight fatigue. They are not associated with the specific activity of the nervous system, that is to say with the explosive discharge of the nervous impulse, any more than they are concerned with the elaboration of secretion in pancreas cells. Key (’16, p. 216) found that the mitochondria are not exhausted in the acinus cells of the pancreas by even prolonged stimulation with secretion and pilocarpin. This independence of mitochondria of the specialized activi- ties of cells is in general accordance with the views which have been expressed by many authors to the effect that they are con- cerned with the basic fundamental processes of metabolism (some hold with oxidations) which all cells possess in common. In fact this is the conclusion to which Cowdry (’14, p. 17) has arrived in the case of the nerve cell. It also falls in line with the generally recognized fact that with starvation and other influences which bring about changes in metabolism, the nervous system is more difficult to alter than any other tissue in the body; as well as with the unsatisfactory results which have usually attended attempts to alter the metab- olism of the nervous system through excessive mental work and other agencies. I refer to the calorimeter experiments of Benedict. Furthermore, these processes of metabolism are, as one would naturally expect, very easily modified in pathological states; MITOCHONDRIA AND NERVOUS ACTIVITY 171 hence the sensitivity of mitochondria to pathological change. Scott (16, p. 250) discovered that the mitochondria are the first of all the cell constituents of the pancreas to become altered in experimental phosphorus poisoning. Moreover, the fact, which is now emerging from the numerous recent pathological studies on mitochondria, that mitochondria in different types of cells respond in much the same way to different varieties of injurious influences, in other words, that there is nothing specific in the reactions of mitochondria to pathological change, is also in ac- cord with the prevalent conception that they take part in a type of activity common to many cells. BIBLIOGRAPHY Busacca, ARCHIMEDE 1915 Sulle modificazioni dei plastosomi, etc. Lab. Anat. Univ. Roma, vol. 18, pp. 217-237. : Cowpry, E. V. 1914 The comparative distribution of mitochondria in spinal ganglion cells of vertebrates. Am. Jour. Anat., vol. 17, pp. 1-29. 1916 The structure of the chromophile cells of the nervous system. Contributions to Embryology, Carnegie Institution of Washington, No. 11. Key, J. ALBERT 1916 On the relation of mitochondria to zymogen granules. Anat. Rec., vol. 10, pp. 215-216. Luna, Emerico 1913 Sulle modificazioni dei plastosomi delle cellule nervose nel trapianto ed in seguito al taglio dei nervi. Anat. Anz., Bd. 44, Pp. 413-415. NicHoutson, Norman Cuive 1916 Morphological and microchemical varia- tions in the mitochondria in the cells of the central nervous system. Am. Jour. Anat., vol. 19, pp. 329-350. Scott, W. J. M. 1916 Experimental mitochondrial changes in the pancreas in phosphorus poisoning. Am. Jour. Anat., vol. 19, pp. 237-253. Tuurtow, M. DeG. 1916 Observations on the mitochondrial content of the cells of the nuclei of the cranial nerves. Anat. Rec., vol. 10, p. 253. AN ANOMALOUS VENA PULMONALIS WITHIN THE LUNG T. STANLEY O’MALLEY From the Anatomical Laboratory of the University of Wisconsin TWO FIGURES Anomalies in the distribution of the Venae pulmonales within the lung, so far as the author has been able to discover, have not been described. The only cases which he has been able to find were extra-pulmonary, and were either variations in the fu- sions of the venous trunks before entering the atrium sinistrum of the heart, or aberrant veins which emptied into the Vena cava superior, the Vena cava inferior or some one of their branches. While making a series of study corrosions of the blood vessels of the lung of the dog and their relation to the bronchial tree, Professor Miller called my attention to an anomaly within the lung which seems to be unique. The lung of the dog consists of four lobes on the right side and of two on the left side. The lobus superior (oral) of the left side is incompletely subdivided into two lobes by a deep notch. The first of these I shall, for convenience of description, designate the apical lobe; the second, the left cardiac lobe. The first hyparterial branch given off by the left bronchus almost immedi- iately divides (figs. 1 and 2) into two main branches which pass, the one to the apical lobe (the apical bronchus), the other to the left cardiac lobe (the ieft cardiac bronchus). In the normal lung the blood is returned from the left lobus superior (fig. 1) by two main venous trunks which are situated ventral to, and slightly removed from, the corresponding branches of the bronchial tree. In the angle between the two main bronchi a small vein is seen (fig. 1) which returns the blood from a portion of the first branch given off by the apical bronchus. This vein passes behind (dorsal to) the bronchial 173 174 T. STANLEY O'MALLEY stem going to the aboral portion of the lobus superior, the left cardiac lobe, and joins the main venous trunk which accom- panies this bronchus. It is necessary to note carefully the ori- gin and position of this small vein, for it apparently plays an important réle in explaining the course of the anomaly. s- s S xs - whe, 27 2 —~ * Os “Ge —_—_ ; LI > Ee 2 Fig. 1 Celloidin corrosion of the lobus superior of the left lung of a dog show- ing the normal distribution of the Venae pulmonales and their relation to the apical bronchus and the left cardiac bronchus. Note the small vein in the angle between the two bronchi. Its connection with the cardiac vein is not shown. In this figure and in figure 2 some of the bronchial branches have been omitted for the sake of clearness. Fig. 2. Celloidin corrosion of the lobus superior of the left lung of a dog in which an anomalous distribution of the Venae pulmonales was found in the apical lobe. Note how the apical vein forms half of a spiral turn about the bronchus and eventually joins the vein coming from the aboral portion of the lobus superior. The first portion of the anomalous vein follows closely the course of the normal vein; but as soon as the two distal radicles have united the vein swings away from the bronchus towards the facies mediastinalis (fig. 2). It then passes dorsal to the Arteria pulmonalis and the bronchus forming half of a spiral turn about these structures, and passing dorsal to the left car- diac bronchus, empties into the vein which accompanies the latter. ANOMALOUS VENA PULMONALIS WITHIN LUNG 175 Only one small branch joins the lateral side of the vein through- out its unusual course. On the mesial side the branches have a normal arrangement. As the vein curves behind the artery and the apical bronchus it is joined by a branch of considerable size which is formed by venous radicles coming from the area that is usually drained by branches which join the lateral surface of the apical vein, when it takes its normal course, and by radicles which normally form the small vein found in the angle between the apical bronchus and the left cardiac bronchus. The trunk formed by the union of these two branches follows, as stated above, the course taken by the small vein found at this point in the normal distribution of the veins. The following explanation of the anomaly seems to be the rational one. During the development of the pulmonary venous system some obstruction to, or atrophy of, the vessels which usually form the apical vein took place. Changes were thus brought about in the capillary bed and secondary trunks were formed which connected with the small vein present in the nor- mal condition in the angle between the apical bronchus and the left cardiac bronchus; as a result this vein became the main trunk and returned all the blood from the apical bronchus. THE ANATOMY OF A DOUBLE PIG, SYNCEPHALUS THORACOPAGUS, WITH ESPECIAL CONSIDER- ATION OF THE GENETIC SIGNIFICANCE OF THE CIRCULATORY APPARATUS EBEN CAREY Department of Histology and Embryology, Creighton Medical College, Omaha, Nebraska TEN FIGURES INTRODUCTION The following case of syncephalus or Janiceps asymmetros, is in- teresting on account of first, the unusual possession of two distinct hearts each contained in a separate pericardium; second, to the subsequent fusion of the aortic arches, of both hearts, with each other, in which there were persistency or degeneration of certain integers to form the anomalous condition of the circulatory apparatus; and third, to the pos- session of two complete cerebro-spinal axes. According to Fischer (’04) there were 50 human and about 80 cases in the domestic animals of the syncephalus class, recorded in teratolog- ical literature. However, I have been able to find only one case of a double pig that shows a close resemblance to the specimen here dissected. Wymann (’61) dissected a syncephalic pig presenting the following characteristics. Extract “A single head in which the nervous system is made up of the right hemisphere of one brain and the left hemisphere of the other with an intermediate compound one. There is a single optic thalamus and striated body, below this the nervous system is double with a distinct cerebellum for each lateral cerebral hemisphere and two spinal cords. The head possesses two lateral ears and a posterior compound one. There are three nostrils on the snout. There are two sets of upper ex- tremities. The body is fused from above downwards to the umbilicus; from the latter point the compound body divides into a right and left body with two distinct sets of inferior extremities. The thyroid, lungs, liver, kidneys, spleen and genitals are double. The heart is compound. The alimentary canal tract is single to the lower one-third of the ileum and from there it is double.”’ 177 THE ANATOMICAL RECORD, VOL. 12, No. 1 17 EBEN CAREY CO The chief differences as will be noted in the following text from the above description by Wymann, will be found in the nervous system and in the heart. He does not describe the remainder of the blood vascular system, so no comparison may be made there. In cases of complete dicephalus, there are found two distinct nery- ous systems in their cephalic aspect at least, and generally two hearts. But in no case in the literature on syncephalus, of the Janiceps asym- metros type, to which the writer had access, either in the domestic animals or in man, was he able to find a specimen recorded possessing two distinct cerebro-spinal axes, and two distinct hearts enclosed in separate pericardia with the anomalous condition of the blood vascular system as here found. A few cases of double monsters in other domestic animals which show some external or internal feature in common with the double pig here studied are described by Dareste (52) in the cat, Gurlt (’32) cat, Mitchell (90-91) chick, McIntosh (’68) cat, Pilcher (’80) pup, Reese (’11) cat. ; The following is a partial list of the cases consulted on syncephalus in man, Braun (’79), Caillé (’89~’90-’91), Debiene et Dutilleul (90), Godson (6870), Hilliard (’80), Hirst and Piersol (’93), Hue (6570), Mayor (’81—’82), McLaurin (’80-’81), von Swiecicki (’87), Walter (’89). The specimen that is the subject of this paper was given to the author in 1914 by Dr. J. S. Foote, professor of histopathology, Creigh- ton Medical College. It was presented to the department of pathology by Drs. Sumney and Hellwig of Omaha seven years ago. From the degree of development of the external body form, the hair, extent of descent of the testicles, palpebral fissures and the foramen ovale, the monster corresponded to the appearance of a fifteen week fetus. The normal period of gestation for the pig is about one hundred and twenty days or roughly speaking seventeen weeks. The specimen was covered entirely by white hair and weighed 500 grams. The right body is longer than the left one as can be verified by reference to the skiagraph, figure 9. The former measured 20 cm., the latter 18 cm. in the crown and rump line. TOPOGRAPHICAL ANATOMY Ventral aspect (fig. 1). It is readily apparent that the heads of the monster were turned laterad, the left head to the left and the right one to the right, and fused to form a single compound head witha ventral compound face and a dorsal rudimentary one. The larger ventral compound face is made up of the right side of one head and the left side of the other. The snout has four nostrils. The two well formed ones belong to the left head and the two rudimentary ones belong to the right. The left and right eyes belong to the left and right heads respectively. ANATOMY OF A DOUBLE PIG 179 The bodies are joined venter to venter. The fusion extends cephalo- caudad to the umbilicus from which point caudad the bodies are dis- tinct and normal. There is but one umbilical cord which contains four arteries, and two veins. This cord is located on a plane mesiad be- tween the two bodies rather than on a plane passing dorso-ventrad through the ventral thoracopagic region. These vessels are distinct Figure 1 for the remainder of the cord left intact with the monster; which was for a length of 6 cm. External examination proved both bodies to be males; this being verified upon dissection. Dorsal aspect (fig. 2). Upon the mesio-cephalic aspect of the fused heads is seen an oval pit devoid of hair which marks the end of the proboscis of a cyclops; this proboscis was formed by the fusion of the 180 EBEN CAREY right eye of the left head and the left eye of the right one. At the base of the skull a pair of small ears of the dorsal face are found close together, which bear the same relationships to the compound skulls as do the eyes. The two dorsal fore limbs are separate integers, the left being the right fore limb of the left body and the right the left fore limb of the right body. The two separate bodies below the bifurcation of the monster are normal. Figure 2 INTERNAL ANATOMY Although the muscles were dissected and studied, no attempt will be made to record here the detailed findings. Respiratory, digestive and genito-urinary systems (fig. 3). There was a ventral and a dorsal pair of lungs. The former possessed a three lobe right and a two lobe left lung; the latter, a three lobed left anda ANATOMY OF A DOUBLE PIG 181 two lobed right lung. Each pair of pulmones is composed of integers heterogeneous in origin. The ventral pair comprises the right and left lung of the right and left bodies respectively; the dorsal, of the left and right lung respectively of the right and left bodies. Each pair of lungs possesses a corresponding trachea which leads cephalad into its respective larynx. Each of the latter was surmounted by an epiglottis. The larynges were at the same level on a mid-dorso- ventral line. The tongue was compound. At the location of the foramen cecum of the right integer there was a pedicle 1 cm. long; on the distal end there was located a globular structure. Upon section this proved to be thyroid glandular tissue. The left thyroid gland was present but the right was absent from its normal locations in the neck. Cephalad the opening into the oesophagus was located between the two larynges. The alimentary tract was single to within 16 em. of the cecum. At the former point the ileum bifurcated, each integer leading to its respec- tive ileo-cecal valve. However, this single tract possessed a much larger ileum than is normally found even in a post-partum pig of two or three weeks of age. Each portion of the now double alimentary tract was normal for its corresponding body. There were two livers the right being larger than the left. These were located considerably more caudad than normally. The common bile-duct from each gall-bladder was joined by the pancreatic duct of the same side 0.25 em. before the entrance into the duodenal wall. There was a normal pancreas for each body. The genito-urinary system was double each entity being normal. The testicles were undescended all four being located respectively in an inguinal canal just internal to the external abdominal ring. CIRCULATORY SYSTEM Ventral aspect (schematic (fig. 4) ). There was one large heart situ- ated ventrad within the thorax in a separate pericardium. Dorsad and to the left was found a heart one-quarter of the size of the ven- tral one. The former evidently had been crowded into the position it occupied in the right portion of the thoracic cavity of the left body by the much larger and more vigorous ventral cor. The smaller one also possessed a separate pericardium. The auricular cavities of each heart were normal in structure. However in both, the auricles were connected by an abnormally enlarged foramen ovale due to the fail- ure of complete development of the septa prima and secunda. The ventricular cavities of both inter-communicated due to the lack of development of the interventricular septa. The large ascending aorta of the ventral heart bifurcated 1.5 em. from its origin into a larger right aortic arch and a smaller left one. At the junction of the left aortic arch with its corresponding ductus arteriosus, the left and right subclavian arteries of the left body are 182 EBEN CAREY epighttis thyroid cartilage ae eee pore ranked Ee eal ee = <= 4 _.. beso phagas Ventral polmones dorsal pulmones * - = = Ee see LY Li common pile dtc, left Ap TT Re Le - JY _tidney Neumpbifuroaiion of umb. ring Figure 3 ANATOMY OF A DOUBLE PIG 183 Pi caaae hoes oarotia left 24, middle cerebral citcle of willis y = ——Pesletior communiceting = ————-besUer soastomosig Jnteriss carotid ectetnel carotid ——‘ommeon cerotig Commos carotid a —Sz7e@°S carotld —— Ved® cAvS superior Tignt aorta———~-———# eu >> forts left Superior Vena cava——__-8 £7 re > Yona cava superior — +--+ a ae ee g GUctus erterissus right subclavian te AW - ; } i f ~ - ) id left SUbclavian f { ——— \\ 5 Sil subclavian Jett a xk florta ri at —+__ subclavian right ductus arteriosus y- Lr -——__43 pulmonary veutra! va = — Yona Cavs jor 4 ral heart re Sade \ventral heart» Mmferior Vena cava _pulmoaery dorsal ra 1 aorta Vene& CSVS inferior __ Sorte epstics- ides hbepatics portal umb. vein — ae ductus venoscs etes venctas umb. vein hypogastric Figure 4 184 EBEN CAREY given off. Coming from the latter arteries their respective vertebral branches are found. The right aortic arch of the ventral heart, Just proximal to its junc- tion with the left aortic arch of the dorsal heart, gives off the right and left subclavian arteries of the right body. From the latter vessels their corresponding vertebrals are derived. One cm. caudad to the junc- tion referred to above, the ductus arteriosus of the dorsal heart joins the descending aorta of the right body. The latter vessel, as will be seen later, belongs genetically to the dorsal heart. From the left aortic arch of the dorsal heart a single vessel arises which is called here the azygos carotid. Four cm. from its origin it bifurcates into the two common carotids. The internal carotids of these latter vessels, go to form the mesial aspects of their respective circles of Willis. The left aspect of the left circle of Willis and the right aspect of the right circle of Willis derive their vascular supply from the in- ternal carotids which are branches of the common carotids of the right aortic arch of the ventral heart. Four hypogastric arteries leave the umbilical ring, one from each of the four internal iliac arteries. There are two umbilical veins which are distributed normally one to each body. The intercostal and lumbar arteries although not depicted in figure 4 are distributed in a normal manner from each descending aorta. There are two spleens. Genetic significance of the circulatory apparatus (figs. 5, 6, 7, 8). In order to simplify the understanding of the means by which such an anomalous condition of the circulation was formed, it will be neces- sary, in the first place, to consider the normal aortic arches of two hearts as presented in figures 5 and 6. The former depicts the normal arches of a heart from the dorsal aspect; the latter those from the ventral view. In the fusion of two embryonic areas or the splitting of a single one to form the monster, the bodies were ultimately brought venter to venter and conjugated; as a consequence the ventral surfaces of each set of arches were brought into apposition. After the fusion and the resultant abnormal alteration of origin of some of the large vessels, we then would have the condition of the circulation as represented in figure 7. Here we find the right and left aortic arches of each heart still persisting. Two common carotids come off abnormally from the right aortic arch of the ventral heart and a single trunk called, as previously stated, the azygos carotid, from the left aortic arch of the dorsal one. It is important to remember that genetically the right common carotid of the right aortic arch of the ventral belongs to the dorsal he: irt; and that the smaller left common carotid of the azygos carotid of the dorsal heart belongs to the ventral one. As schematics uly shown, the subclavian arteries are still branches of their respective right and left. arches. In figure 8, we find the complete degeneration of the proximal portion of the right aortic arch of the dorsal heart; the subclavian ar- tery of this vessel is now a trunk of the descending aorta of the ventral ANATOMY OF A DOUBLE PIG external carotid internal carotid Common carotid aorta left left vertebral bleft sobclavian ductus arteriosus —______Pulmonary left = sorte Pulmonary Pulmonary right —_—___ ZOTtS Tight vertebral TIERS subclavian common carotid left sorte left right sorta Pulmonary vyantral &zygos carotid right vertebral left vertebral — tuctas arterioens ee eh left vertebral right subclavian left subclavian left subclavian ———-t ight sabciaviay ductus arteriosus ————-Tight vertebral dorsal pulmonary — left aorta — > ied sorta $ left left right aorta left vertebral —— right vertebral tight sudclavian left acrta—— left subdclavian Guctus arte dorsal PUlmonary left aorta left body 185 186 EBEN CAREY heart and supplies the right extremity of the left body. The left sub- clavian of the left aortic arch of the dorsal heart is abnormally a branch of the first part of the descending aorta of the ventral heart and fur- nishes the blood supply to the left extremity of the right body. From the above embryological consideration it is seen that the larger ventral heart belongs genetically to the left body whereas the smaller dorsal heart belongs to the right body. However, the dorsal heart has been rotated and crowded from its earlier position into the right side of the thoracic cavity of the left body by the larger and more vigorous ventral heart. The ventral surface of the dorsal heart is now directed ventrad, in reference to the double monster, instead of dorsad. As a consequence the relationship of its great vessels is considerably altered. The right and left aortic arches of the ventral heart have persisted, the right being considerably the larger of the two as previously pointed out. The right by joining the descending aorta of the dorsal heart form a good size trunk, slightly larger than the left one. This is due to the fact that both hearts contribute to the descending aorta of the right body and only the smaller left aortic arch and its corresponding ductus arteriosus of the ventral heart go to form the descending aorta of the left body. It is therefore clearly seen that the right and larger body receives the greater blood supply, “due to the influences which have brought about a more abundant growth of capillaries,’ Evans (712). Developmentally the vessels of the dorsal heart supply the right side of the left body and the left side of the right body whereas those of the ventral one, supply the left side of the left body and the right side of the right body. In other words, the larger ventral heart sup- plies the ventral aspect of the syncephalo-thoracopagic regions, of the double monster and the smaller dorsal heart genetically contributes the vessels distributed to the dorsal aspect. In the lumbar, pel- vic and caudal extremities of each body, the vessels are normally distributed. The same conditions that governed the relationship of the areas supplied by the arteries of the ventral and dorsal heart ruled over the veins draining those regions. However, the entrance of the veins can be traced to their respective hearts, whereas on the. other hand, the genetic relationship at first is somewhat obscure and blotted out in regard to the arteries. Each heart possesses two superior venae cavae representing the persistent anterior cardinal veins which emptied separately at the right and left lateral poles of the sinus venosus. Although figures 5 and 6 are schematically represented as normal arches, it is by no means the intention of the writer to convey the impression that two absolutely normal aortic arches, developed to the stage as here represented, were necessarily at one time existent, or that fusion took place at this late period in development. The establish- ment of this anomalous circulatory apparatus was no doubt forecast in the fused capillary net-work which was the anlage of the abnormal ANATOMY OF A DOUBLE PIG 187 condition of the arches as shown in figure 8. The early indifferent endothelial vascular network of each heart fused and then the elabora- tion of the main trunks as depicted in figure 8 represent a physiologi- cal adaptation of this fused network to the demands of the circulation in the monster. The fortuitous location of certain channels of the net with regard to the aorta of either heart determined the enlargement and use of these paths to form the normal and abnormal vessels. It was merely for the sake of simplicity to emphasize the venter to venter apposition and fusion of the anlage of each set of aortic arches, that two developed normal sets are used in diagrams, instead of the com- plex antecedent network of each. As a consequence correct sequence of development of the vessels was sacrificed for simplicity. THE SKELETON The posterior extremities, pelvis and lumbar vertebrae were nor- mal for each body. In the dorsal region of the left vertebral column, from the third to the ninth dorsal vertebrae, kyphosis is presented. This is undoubtedly the cause for the shorter length of the left body. The thorax was compound, the sternum and ventral ends of the ribs of each side were so reflected and fused with their corresponding parts of the other as to form one large thoracic cavity bounded by fifty-six ribs; fourteen on the lateral aspect of each body. Each sternum, dorsal and ventral, has a double origin as is apparent from the reflection considered above. Only the upper seven ribs were at- tached to either sternum. The remaining seven were absolutely free, having neither cartilagenous nor bony connections at their distal ends. The thoracic appendages were normal in structure. How- ever, the ventral pair were better developed than the dorsal pair. The spinous processes of the cervical vertebrae were directed laterad instead of dorsad which fact may readily be verified by a reference to the skiagraph, figure 9. The skull is compound. This is apparent for each vertebral column has its separate occipital attachment. In the fusion the interior of the compound cranium was left double. The right tem- poral, parietal, frontal, nasal, parietal maxillary and lateral aspect of the occipital bones of the left skull fused with the corresponding parts of the right one, to form an attenuated partition between the left and the right encephala. This line of fusion may be seen from the exterior extending through the oval depression marking the point of emission of the proboscis of the cyclops, marked 2, figure 10. The two opposed orbits were fused to form the condition of synopthalmia. The par- tition did not leave the normal amount of space for the development of each brain; this is apparent by the fact that the mesial hemisphere of each encephalon is smaller than the lateralone. The adjacent walls of the mesial hemispheres present parallel surfaces instead of being normally convex. 188 EBEN CAREY Figure 9 ANATOMY OF A DOUBLE PIG 189 Fig. 10 Cerebro-spinal axes. Dorsal aspect. 1, olfactory bulb; 2, proboscis / > of the cyclops; 3, right cerebrum; 4, cerebellum; 5, medulla oblongata; 6, cervical swelling; 7, lumbar swelling; 8, cauda aquina. 190 EBEN CAREY CEREBRO-SPINAL AXES There are two encephala in the double compound cavity. Each brain and stem is distinct, there being no fusion. However, the right optic nerve of the left encephalon and the left one of the right enceph- alon do fuse to form a compound trunk. This terminated in a small amorphous structure which represents the fused eyes of the cyclops, marked 2, figure 10, asreferred to above. The optic nerve of the right lateral normal eye belongs to the right encephalon; that for the left lateral normal eye is derived from the left encephalon. Neither is seen in figure 10, due to the fact that the cerebrospinal axes were so placed that the center of focus would fall on the synophthamia. Each encephalon as a result obscured its corresponding lateral eye from view. The remainder of the cranial nerves were distinct and separate. -Except for the optic fusion as here considered, there is no other connection between the cerebro-spinal axes. The effect of kyphosis of the dorsal vertebrae on the contained cord is readily apparent in figure 10. The marked loop to the left in the left spinal cord in the upper dorsal region is clearly seen. In actual measurement both cords have the same length. The shortening is due to the loop caused by the kyphosis circle of Willis. Each heart contributes blood to each encephalon as was seen above. The dorsal heart furnishes the blood supply to the mesial aspects of each circle of Willis, whereas, the ventral heart furnishes the blood to the lateral aspect of each circle of Willis. In conclusion, I wish to express my sincere thanks to Dr. Foote for the specimen here recorded and to Dr. Tyler, professor of Roent- genology of Creighton Medical College, for the skiagraph of the skeleton, figure 9. ANATOMY OF A DOUBLE PIG 191 LITERATURE CITED Braun, E. 1879 Ein Fall von Doppelmissbildung. Wien. Med. Presse, 20, 275-277. Caitit, A. 1889 Janiceps asymmetros. Arch. Pediat. Phila., 1889, 6, 822. 1888-89-90 Janiceps asymmetros. Trans. Amer. Pediat. Soc., Phila., 1, 70. DarestE, C. 1888-89-90 Memoire sur un chat iléadelphe a téte monstreuse. Annales d. Sciences naturelles: Zoologie, vol. 18, pp. 81-94. DEBIENE ET DutILLEUL 1890 Contribution a l’étude des monstres doubles du genre synote. Arch de Physiol. norm. et Path, Paris, 5, s., ii, 45-58. Evans, H. C. 1912 Keibell and Mall Human Embryology, vol. 11, p. 581. Fisoer 1904 Teratology Ref. Hdbk. Med. Sc., new ed., vol. 7. Gopson, C. 1886 Monster ‘Double Syncephalien,”’ dissection by D’A. Power. Trans. Obs. Soc. London, 27, 68-70. Gurut, E. F. 1831-32 Lehrbuch der pathologischen Anatomie der Haussauge- thiere, Berlin. Hitirarp 1880 One-headed twin monster. Obs. Journ. of Gr. Brit., London, 8, 91. Hirst anpd Piersot 1893 Human monstrosities. Philadelphia, vol. 4, pp. 170-172. Hur, F. 1889 Présentation d’un monstre double antositaire syncéphalien synote. Bull. Soc. de Med. Rouen, 2, s., ii, 65-67. Mayor, A. 1882 Contribution a l’étude des monstres doubles; des monstres du genre janiceps. Arch. de Physiol. norm. et Path., 2, s., 9, 127-161. 1882 Note sur un monstre genre Janiceps. Progres Med., Paris, 10, 224-244. MclInrosu 1868 Notes on the structure of a monstrous kitten. Journ. of Anat. and Physiol, 366-373. McLaurin, H. N. 1881 Twin monster, Syncephalus. Trans. Obs. Soc. of London, xxii, 155. Mircnett 1890-91 On a double-chick embryo. Journ. of Anat. and Physiol., vol. 25, pp. 316-324. Pitcuer, L. S. 1880 Double monsters. Annals of the Anatomical and Sur- gical Soc. of Brooklyn, vol. 2, No. 1, pp. 19-37. Reese, A. M. 1911 The anatomy of a double cat. Anat. Rec., vol. 5, No. 8, pp. 383-390. \on Swiecickt 1887 Die Geburt eines Janiceps. Centralbt. f. Gynak., Leip- zig, 11, 845-847. Water, W. H. 1889 Double monstrosity. Brit. Med. Journ., London, vol. 1, p. 1405. WyMANN 1858-59 Dissection of a double pig. Boston Med. and Surg. J., 64, 535. A DEVICE TO INCREASE THE EFFICIENCY AND EASE OF MANIPULATION OF THE FINE ADJUST- MENT OF THE MICROSCOPE ROBERT T. HANCE Zoological Laboratory, University of Pennsylvania ONE FIGURE The use of high powered microscopic lenses requires the hand to be constantly on the fine adjustment. When the study is continued for several hours at a time the hand and arm doing the focusing, even though the latter is supported at the elbow on the table or chair arm, become extremely fatigued. To obviate this strain the apparatus fig- ured below was devised. Although the role it was originally in- tended to fill was merely to facilitate the use of the fine adjustment, it has, since its installment, not only been of aid in this way but has greatly diminished several annoying features that are frequently con- nected with high power work. For this reason it seemed worth while to bring the attachment to the attention of microscopists. The device as figured is for use with microscopes which have the more common ‘top’ fine adjustment although it would be a simple matter to construct a similar apparatus for use» with the side wheel type. No dimensions are given, as these must be controlled by the height of the particular microscope the attachment is intended to be used with. The illustration, which is a side view, is just one-half the size of the one I have in use. The entire apparatus is made of wood and it is simpler to use dowel sticks of various diameters for the pillar (5), the arm (6), and the shaft (3), although other material could be used satisfactorily. The focusing wheel (/) is a dise cut from lumber about one-eighth of an inch thick. It is of advantage to have this wheel large (two to two and one-half inches) and thin, so that it may be easily gripped and turned with either the thumb or fingers. The edge of this disc is rounded and notches are filed into it at intervals of about one-quarter of an inch to prevent the fingers gripping it from slipping. The grooved wheel (2) is made by gluing together two discs about one inch in diameter (cut from thin lumber) whose adjacent rims have been beveled. It is well to have the channel or groove rather deep so as to prevent the belt from jumping when the microscope is inclined. Care, of course, must be taken to center the hole in the arm (6) through which the shaft (3) passes and the socket in the base (4) into which ‘the end of the shaft slips. Smoothness of action is obtained by soaking that part of the shaft that is in contact with the arm, and the hole in the arm through which the shaft passes, with paraffin. The socket in the base is filled with paraffin so that the shaft is resting in a 193 tightly packed bed of this material. I am indebted to Dr. C. E. McClung for suggesting this method of lubrication, which results in a very soft, smooth movement. In use the apparatus is screwed to the table (in my case to the drawing board on which the microscope rests) with the pillar (5) away from the observer. A belt composed of string which will not stretch (such as braided fish line) is passed around the pulley (2) and the focusing wheel of the microscope. The microscope may be used in any position. The focusing arm lies flat on the arm of the chair or the table and the focusing wheel (1) is within easy access. Since the pulley (2) is made smaller than the fine adjustment wheel of the microscope the sensitiveness of focus is increased. The deli- cate focusing of high powers has been found to be much steadier with this attachment. The annoying feature of the field under ob- servation being a trifle displaced while drawing, due to the Hand on the focusing wheel jarring the microscope (however little), is largely eliminated with the use of the device described as it is now unneces- sary to directly touch any part of the microscope. 194 ON THE MECHANISM OF MORPHOLOGICAL DIFFER- ENTIATION IN THE NERVOUS SYSTEM ‘II. THE RELATION BETWEEN COMPRESSION AND THE DEVELOP- MENT OF A SERIES OF VESICLES. OTTO C. GLASER From the Zoological Laboratory of the University of Michigan EIGHT TEXT FIGURES AND THREE PLATES I. INTRODUCTION The simple neural tube mentioned so frequently in embryo- logical literature is really no more than a convenient fiction. Practically it cannot be found at any stage of development. The mere differences in the amounts of tissue in the anterior and posterior ends of the neural plate alone preclude this possibility, but in addition, long before the separation of the plate from its extra-neural ectoderm is complete, the developing ‘tube’ pre- figures a serial differentiation that culminates in a succession of vesicles, within limits, highly constant for the vertebrate ner- vous system, and, as one of its basic attributes, calling for explanation. The stages in embryogenesis upon which a study of this prob- lem must be based, are necessarily as commonplace as those dealt with in my first paper! yet despite their familiarity with this period of development, embryologists do not tell us why the nervous system differentiates a series of vesicles, and par- ticularly why there are five such fundamental divisions in its anterior end. Many do not even ask the necessary questions although His, over forty years ago, considered the subject as legitimately within the province of the student of development. 1On the Mechanism of Morphological Differentiation in the Nervous System. l. The Transformation of a Neural Plate into a Neural Tube. Anat. Rec., vol. 8, pp. 525-551, 1914. 195 THE ANATOMICAL RECORD, VOL. 12, No. 2, MARCH, 1917 196 OTTO C. GLASER In fact His? himself attempted to explain serial differentiation. The analysis, in Unsere Korperform, begins with the ‘closed neural tube,’ whose anterior end, as indicated in figure 1, is larger than its posterior, and is characterized by three en- largements—the precursors of the five secondary brain vesicles. In a later stage, figure 2, His illustrates the relative positions of the five secondary vesicles in the flexed state. The curve, typical of cranial flexure, is divided into a ‘Briickenkriimmung,’ involving the fifth and fourth vesicles; a ‘“Mittelwélbung,’ in- volving the mid brain; and the ‘Hackenkriimmung,’ involving the second and first. At the extreme anterior point of the first vesicle is the ‘Trichterfortsatz’ Tr. His then attempts to show how this form can be derived from that given in figure 1. The brain, according to the argument, begins as a tube whose comparatively large lumen is enclosed by moderately elastic walls. These conditions are identical with those given in a piece of rubber tubing. If the edge of a rubber tube is made fast by threads in the manner indicated in figure 3 longitudinal compression will pro- Fig. 1 After His. Chick embryo of the second day. H, brain, with three vesicles indicated; Ag, optic vesicle; W, Wolffian ridge; Uw, somites; Ump, un- segmented mesodermal bands; Ami, head fold of the Amnion; Am2, lateral folds of the Amnion; Mp, open portion of the neural tube. The heart is indi- cated by means of dotted lines. Enlarged 20 X. Fig. 2 After His. Brain of chick embryo of the third day. Ag, optic ves- icle; Vh, forebrain; Tr, “Trichterfortsatz;’ Zh, interbrain; Mh, midbrain; Hh, hindbrain; R, location of fourth ventricle; Br, ‘Briickenkriimmung;’ Nh, after- brain; Gh, auditory vesicle. Enlarged 30 X. Fig. 3 After His. Rubber tube whose upper end has been drawn backwards by means of a thread. Fig. 4 After His. For explanation of legends see figure 2. The dotted line indicates the anterior limits of the foregut. Fig. 5 Sagittal section through chick-head about 38 hours old, showing the relations of foregut, floor of the second brain vesicle, and the anterior end of the notochord. Fig. 6 After His. Upper figure, head of embryonic pike; middle, of trout, and lower, dorsal view of same. Legends as in figure 2. Rg, olfactory pit. ? Wilhelm His: Unsere Koérperform und das Physiologische Problem ihrer Mntstehung. F. C. W. Vogel, Leipzig, 1874. Achter Brief, pp. 93-104. DIFFERENTIATION IN THE NERVOUS SYSTEM 197 198 OTTO GC. GLASER duce sn abrupt bend just behind the level of fixation, and there will appear in this region a pair of lateral swellings, whereas on one side—either the upper or the lower, as the case may be,—a sharp groove, deepest in the median line, shallower toward the edges of the sidewise expansions, will cross the tube in a trans- verse curve with concave face forward. His points out how exactly this form duplicates the ventral surface of the second vesicle in the stage of development depicted in figure 1. The suggestion that identity of form in the two sets of cases 1s the result of identity in the mechanical conditions under which they were produced is very much strengthened, as His empha- sized, by the union “welche durch den Axenstrang und spiter durch die aus ihm entstandene chorda dorsalis zwischen der Medullarplatte und dem Darmdriisenblatte, lings der Mittel- linie unterhalten wird.’” This union later undergoes the following modifications: first the entoderm separates from the chord, und, viel spiiter, diese vom Medullarrohre . . . . Am innigsten ist die Verbindung durch zwischengelagerte Masse zwischen dem Urspriinglich vordersten Rande der Medullarplatte und vom vorderen Ende des Vorderdarmes. Die Verbindung ist hier eine so innige, dass, wenn in sehr spiiter Zeit der Vorderdarm vom Gehirn sich trennt, die Trennung nicht im Verbindungsstiicke geschieht, sondern in der Con- tinuitit des Vorderdarmes selbst. Ein kleines stiick von diesem bleibt als Vorderer Lappen der Hypophysis in dauernder Verbindung mit dem Gehirn.! His continues, Es wichst aber das Medullarrohr, und speciell das Gehirn rascher in die linge als der Vorderdarm; da es nicht zu einer Trennung beider Theile kommt, so muss der lingere Theil sich Kriimmen, und miissen ferner die unmittelbaren Folgen der Zerrung in den mit einander verbundenen Strecken des Vorderdarms sowohl, als des Medullarrohres mi Tage treten. Beides trifft in sehr prignanter Weise ein, nicht allein erhebt sich das Medullarrohr iiber dem Vorderdarm in wasch- endem Bogen, sondern es ziehen sich an beiden Theilen die Verbun- denen Enden trichterf6érmig aus, wir bekommen auf die Weise am 3 Loe cit., p. 99. * Loc. cit., p. 100. DIFFERENTIATION IN THE NERVOUS SYSTEM 199 Gehirn den oben betrachteten Trichterfortsatz (fig. 2), am Vordarm die . . . . bekannte sog. Rathke’sche Tasche.° The union of the ‘trichterfortsatz’ with Rathke’s pouch is, under the conditions of growth, mechanically the exact equiva- lent of the piece of string in figure 3. The striking similarity of the forms produced in the two cases scarcely requires com- ment and may almost be considered proof of the correctness of the assumptions. One factor, not emphasized by His, however, is the notochord. Given the fusion between ‘Trichterfortsatz’ and Rathke’s pouch, flexure resulting from the faster growth of the nervous system would be accentuated by the notochord whose anterior end would play the réle of a fulerum about which the curvature would take place. But His’ account is not free from anachronisms, nor will it bear a too close scrutiny from the standpoint of comparative anatomy and embryology. The final conclusion which he wishes drawn, “von der Abhingigkeit . . . . in welcher die Gehirngliederung von den auftretenden Longitudinalbiegungen des Organs steht,’’® does not follow. If this were correct, sharp- ness in the demarcation of the vesicles from one another should vary directly with the degree of cranial flexure in individual as well as comparative ontogeny. This, for the first case is not strictly true, and, for the second, scarcely at all, since the vesicles are distinct before cranial flexure, and forms exhibiting a high degree of flexure have their vesicles no more sharply delimited than those in which the bending of the embryonic head is never very marked. I have no reason to doubt that His was correct with respect to the origin of both the flexure, and the lateral expansions, or optic lobes, but the differentiation of the vesicles themselves cannot be explained as the result of flexure. II. ON THE EFFECTS OF DIFFERENTIAL GROWTH It is clear that His considered differential growth the cause of flexure, and flexure the cause of vesiculation. Since, however, only the first vesicle with its ventral groove and lateral expan- 5 Loc. cit., p. 100. § Loc. cit., p. 104. 200 OTTO C. GLASER sions can be accounted for by flexure, the case for this factor is hardly made out. From his manipulation of rubber models, forced in various ways to simulate special regions of the nervous system, as well as from the general tenor of the argument, it is likely that His would not have denied that differential growth, in the early stages of flexure, results In some sort of compression in the longitudinal axis. Be this as it may, it is certain that he at- tributed to this factor not only flexure, but also the changes in relative position which the vesicles undergo during later stages of development. Concerning the head of the embryonic pike, figure 6, he writes: ‘‘ Durch die wachsende Zusammenschiebung der Theile ist der hintere Hirnabschnitt, oder das Nachhirn unter die davor liegende Anlage des Kleinhirns, und diese unter diejenige des Mittelhirns geschoben worden.’’? lll. HIS’ DEMONSTRATION OF DIFFERENTIAL GROWTH His attempted to demonstrate the differential growth of the nervous system on chick embryos ranging roughly from the twenty-fourth to the ninety-sixth hour of incubation.’ The method consisted in comparing at four levels, those of the eye, the ‘Gehirnblase,’ and the first pair of somites, the areas in transverse section of the ‘neural tube’—thought of as spread out flat, and the ectoderm, measured from the median axis to the point of fusion with the lateral muscle plate. On this basis, he was able to convince himself that the nervous system actu- ally grows at a faster rate than the tissues with which it was compared. Although this method may be capable of demonstrating the point, it is certainly not able to show that differential growth in width is accompanied by differential growth in length. The 7 Loc. cit., p. 103. * According to his own statements, the youngest stage used was of the sec- ond day, but his figure with its nine pairs of somites hardly bears this out. See figure 1 of the present paper. DIFFERENTIATION IN THE NERVOUS SYSTEM 201 association of the two is of course very probable, but by no means necessary. Differential growth in length can be determined directly only by measurements in the long axis, and if the values so secured have a certain sense and magnitude, they may be made the basis for inferences concerning compression in that axis. 1V. METHOD OF DETERMINING COMPRESSION IN THE LONG AXIS In order to discover the presence or absence of compression in the long axis it is necessary to find at least one measurable relation which differential growth either changes or brings about. This relation must be longitudinally effective, widely applicable, in magnitude independent of absolute measurements, and finally, capable of exact expression. Many attempts were made to satisfy these conditions before it was found that the necessary data, accuracy, and reliability were obtainable in a very simple way. With the camera lucida, optical sections of embryonic chick heads, at convenient magnification and a focus giving maximal outlines, were carefully traced. About these outlines I then erected the system of lines indicated in figure 7. The base of this system is a line tangential to the anterior face of the first pair of somites. Upon this base perpendiculars, themselves tangential to the sides of the head at its widest level, were erected, and finally, parallel with the somitic base line, a tangent to the anterior edge of the head. In this way the entire cephalic region is included within the area of a rectangle. The purpose of these preliminaries is to get a measure of the length of the head. Without the rectangle it would be easy enough to find an anterior point of reference, but it is never pos- sible to tell exactly where the posterior limit of the head is. In fact in the early stages this grades so insensibly into the body that any posterior limit permitting comparison between various embryos and different stages, of necessity has to be chosen arbi- trarily. If then an arbitrary point is imposed by the conditions of the case, it is best to choose one whose relations to the rest 202 OTTO C. GLASER of the embryo are not only significant, but also likely to present the greatest relative constancy. The somites possess these qualifications more than any other structures, and so I took the longer sides of the cephalic rectangle which rests upon the first pair, not as the, but as a measure of head length.’ Since we de- sire a measure of differences in the rate of lnear growth be- Fig. 7 Diagram to illustrate the cephalic rectangle. The line tangential to the anterior face of the first pair of somites, which are indicated in solid black, is the somitic base line. The perpendiculars erected upon this line are tangential to the head at its widest points. The head-length was arbitrarily determined along these perpendiculars as the distance between the somitice base line and the parallel tangent to the anterior surface of the head. ’To carry out these and the subsequent measurements on live embryos pre- sents, for the present, insuperable difficulties. Fixed material, no doubt, dif- fers in absolute values from the living, and it is possible that the relations be- tween measurements which I shall discuss, are also not the same. However, absolute \ i! les nee d not coneern us at all in the present connection, and if the relative valu ipon which my argument rests were seriously changed, | should hardly expect the constancy in sense which they exhibit. DIFFERENTIATION IN THE NERVOUS SYSTEM 203 tween the neural mass and the head, the simplest method is to compare the length of the head with the length of the nervous system, in various stages of development, measured from the somitic tangent as a base. Provided the head of the embryo exhibits no serious irregularities, its length may be determined arbitrarily in the manner indicated above, but, on account of the vesicles, the length of the ‘brain’ cannot be given by the cephalic rectangle. Several methods involving the volume of the brain were tried but discarded in favor of one which in addition to simplicity, is capable of giving exactly and directly the very information that is wanted. All that is necessary is to determine by means of a map-measurer the perimeter of the nervous system beginning at its intersection with the somitiec tangent on one side and end- ing at the corresponding point on the other. The length of the nervous system will be half this perimeter.'° V. THE NEURO-CEPHALIC QUOTIENT With the aid of these measurements it is possible to express in the form of a fraction the relation in each stage between the length of the head and that of the nervous system. The frac- tion chosen is derived by dividing half the neural perimeter into the head length and tells us how many units of head length, within the limits of the cephalic rectangle, are available for every unit of length in the nervous system. This fraction I shall call the Neuro-Cephalic Quotient. Very few embryos approach the ideals of rectitude and sym- metry depicted in text-books and wall-charts. Out of the en- tire collection with which I have worked I have been able to find only eight justly to be characterized as diagrammatic. The outlines of these are reproduced in plate 1, figures A, B, C, D, E, F, G, and H, whereas the number of somites, and the corresponding neuro-cephalic quotients are given in table 1. 10 Since this method is applicable to the nervous system it might be asked why J did not apply it also to the head instead of relying on the long sides of the cephalic rectangle. The answer is, because the perimeter of the head in early stages is distinct only in the anterior region. 204 OTTO C. GLASER On account of the small number of cases the scientific value of this particular table is negligible. Nevertheless it suggests cer- tain problems and questions which must be dealt with before we can arrive at a just estimate of the significance of the quotient. The values, taken for the time being as they stand, seem to indicate that when diagrammatic embryos are arranged in series according to somites—or, in other words, according to age and degree of development, the size of the quotient will be found to rary inversely with the number of somitic units. This, if cor- rect, means that as development goes on the amount of head length into which a given length of nervous system must fit, decreases progressively. TABLE 1 EMBRYO eee QUOTIENT ) ES A ee I Ree Pare Ws AS haha oe Cs Ae RO 2 1.485 Bee i sane sine os ae Nissen ie oe ge ee 8 0.902 Oe Bie Se xcs shed oe cles ee 9 0.877 Der tense sie 0 Steak faa 2 eet ee 9 0.837 | ee ee eee ree Mey en eS os 10 0.820 We ev eine Sa bed Oh, Reo os eee ee 10 0.820 Wie, caciaya Sm «ales efeveie, sty ates Ree eee ne 10 0.846 Eolas oo 55x Gt Sh Aig eee ap eee 11 0.772 Given this condition, differentiations of some sort, spreading, collapse, telescoping, or vesiculation, are to be expected. Leav- ing aside, for the moment, the question why vesiculation pre- rails, let me explain why I choose to consider first these rare diagrammatic forms. The reason is very simple: because in these, the presence of a high degree of symmetry indicates that the developmental processes in general have been in nearly perfect balance, and have given a result relatively or quite free from complications calling for special explanations. Briefly, these forms are thé simplest available. Granted the advantage of simplicity—we must ask why a well balanced development gives rise to the obvious discrepancies which the table exhibits and furthermore why it is itself so rare. The effect of error introduced by the various technical methods employed in preparing the embryos for study cannot be denied, DIFFERENTIATION IN THE NERVOUS SYSTEM 205 nor can I claim for my measurements a maximal degree of ac- curacy. Nevertheless I see no reason for suspecting greater errors than inhere in embryological and biometric work in gen- eral. For reasons which I shall attempt to make clear, { be- lieve that the major discrepancies inhere in the material itself. Referring to the table, the general sense of the values is very obvious. There is, however, no absolute proportionality be- tween somitic increase and the shrinking quotient. Further- more, embryos like C and D, each with nine somites, have quotients separated by a wider margin than C and G. A series arranged according to somites, even in diagrammatic forms, does not coincide absolutely with a series based on quotients. That there is a general coincidence, however, will hardly be denied. It was the desire to find embryos in which just this sort of discrepancy could be expected to assert itself in minimal degree that lead me to select the most diagrammatic forms for separate treatment. But even in these complete elimination of discords is impossible. If now the lack of harmony between theory and practice is to be sought in the embryo itself rather than in the methods by which it was handled, we must analyze our material in an attempt to get at the real explanation. Such explanation will be found, I think, when we realize the true nature of the developmental process itself. To be alive is to solve a constellation of interlocking problems in equilibration. In the adult, departures from balance occur within compara- tively restricted compass, and, being for the most part quickly reversed, result in few or relatively unimportant morphogenetic changes;!! in the embryo, on the contrary, the excursions are often very wide. Indeed it is hardly too much to say that a developing organism ‘blunders’ from one crisis to another, until gradually, by the narrowing of its ‘horizon,’ it reaches that state of relative stability which is characteristic of the adult. Nothing that happens in the fully developed organism can be compared with the multiplicity and complexity of the immediate and remote adjustments consequent upon the differentiation of 1 See Glaser, The Basis of Individuality in Organisms. Science, vol. 44, pp. 219-224. 206 OTTO C. GLASER the germ layers From the dynamic standpoint, development might be defined as the symptom of an organic instability in which departures from exact balance occur within limits so wide as to escape fatality by only a narrow margin. This granted, it follows that the sum of opportunities within the developing system is exceedingly great. Although unques- tionably exact and theoretically predictable in all its details, embryogenesis, within the boundaries of what is ‘normal,’ never- theless, varies tremendously. It need occasion no surprise then if we find differences in the several tissue-maneouvres, or in the exact time of onset of this, that, or the other process. Of several embryos which might be expected to exhibit identical conditions in all respects, one may lead or lag in the morphogenetics of its nervous system, a second in that of its somites, and another in its circulatory equipment. In fact the early discrepancies or temporary misfits of development may at any instant simulate disorganization. By this it is not intended to suggest that ‘normal’ embryogenesis is strictly a matter of chance, but only that its ‘administration’ appears relatively loose. To us this looseness is emphasized subjectively because we remain ignorant of so large a share of the elements underlying the process. We may be sure, however, that the minor errors of development sooner or later receive adequate correction for the end results of embryogenesis are precise and give an organism which actu- ally describes the genetic constitution of its ancestors." Taking development as it is, our quotient, to have significance, must be applicable to a wide range of cases, which, no matter how they may deviate from the diagrammatic, nevertheless cannot be considered otherwise than normal. Within this range, if our preliminary test is to be trusted, we should find the same general relations exhibited by the ideal embryos. However, since the measurements upon which the quotient rests, them- selves bear no obviously immediate relation to the factors upon which the production of somites depends, we should not expect absolute correspondence between somitic increase and a falling 12 See Note 11. DIFFERENTIATION IN THE NERVOUS SYSTEM 207 quotient. All that we are entitled to expect is that the quotient in general will vary inversely with the number of somites. If the quotient is a true indicator of compression in the longi- tudinal axis, and if compression Is related causally to serial differ- entiation, we are also entitled to expect some relation between this differentiation and the quotient. Here again, we should not set our minds on absolute correspondence, for quite apart from the difficulty of exact determinations in this connection, the amount of differentiation which a given degree of compres- sion calls forth depends on many things. Age is one; what dif- ferentiations had taken place before a particular degree of compression was reached, is a second; the thickness and mechani- cal properties of the nervous system are a third; the rate at which the differentiations are produced is a fourth; the exact axis of maximal compression is a fifth; and no doubt there are many others. There is, however, no accurate way, particularly in the more complicated cases, of expressing the degree of differ- entiation. Furthermore the attempt to do so would necessitate also the consideration of that portion of the system which lies posterior to the somitic base line of the cephalic rectangle. Nevertheless, little serial differentiation should be associated with a large, and the reverse with a small quotient. Correspondences between quotient and somites on the one hand, and quotients and differentiation on the other, do not complete the list of what we may expect. A third matter— that of aberrancies—must receive consideration. From our present standpoint, the aberrancies theoretically to be considered are those in which the nervous system is either underdifferentiated or has erred or has been forced to err seriously in the opposite direction. In the first case we should expect relatively high, in the second, relatively low, quotients. As a matter of fact, I have not found enough cases of undifferentia- tion to feel warranted in giving them special prominence, but overdifferentiation is common. In fact, in one of its varieties, it is too common to leave much doubt that the resulting forms must be classified as normal. This type, characterized by the partial telescoping of the first and second vesicles exceeds in 208 OTTO C. GLASER frequency the known percentage of faulty hatchings or total failures by so large a margin as to suggest very strikingly indeed the likelihood that many nervous systems during some stage of their development exhibit temporarily, at least, too much dif- ferentiation. Cases in point are illustrated on plate 2, embryos iJ, Bee, eA: To what extent now, our several expectations are fulfilled can be seen by comparing the figures of the forms illustrated in plates 1 and 2, with the corresponding values given in table 2 in which the embryos are arranged in a series beginning with the highest and ending with the lowest quotient. The omission from this table of 12 and 14 somite embryos is due to the fact that only one of each kind was available. The only other grounds of elimination were asymmetry or a degree of aberrancy which could have but one interpretation. Such em- bryos with somites and quotients indicated are referred to in table 3 and illustrated in plate 3. If we accept table 2 as indicative of the norm for the various stages dealt with, the quotients in table 3 become significant. As can be seen by reference to plate 3, the embryos now under consideration fall into two groups—in one of these, including U, W, and X, the embryo is asymmetrical, in the other, T, V, Y, and Z, the embryos are overdifferentiated. With respect to Z, nothing definite can be said, since table 2 does not contain the quotient normal for the 27 somite stage. Embryos T, V, and Y, however, all have quotients too low for their respective ages. In other words the compression to which the nervous system is subjected is higher than normal and has resulted in too much differentiation. The asymmetrical forms, U, W, and X, suggest that if a cer- tain degree of compression fails to set in, or is not properly di- rected, the aberrant condition will be reflected in an asymmetrical distribution of the neural mass. CONCLUSION A comparative study of the illustrations in the plates and the values given in tables 1, 2, and 3 may, I think, be justly sum- marized by saying that our expectations in general have been DIFFERENTIATION IN THE NERVOUS SYSTEM 209 TABLE 2 Normal embryos QUOTIENT SOMITES REMARKS 1.890 3 12672 2 1.485 2 1.313 4 1.132 3 1.0475 5 1.017 5 1.012 4} 1.0095 5 0.968 6 0.952 6 0.949 6 0.925 8 0.920 7 0.911 10 0.903 7 0.902 8 0.9009 9 0.887 9 0.877 9 0.871 10 0.863 9 0.851 10 0.849 10 0.847 10 0.846 10 0.842 10 0.842 10 0.837 9 0.820 10 0.820 10 0.818 1h 0.812 11 0.811 10 0.800 11 0.772 11 0.737 13 Telescoped forms Plate 2 I 0.664 13 Telescoped forms Plate 2 J 0.649 15 Telescoped forms Plate 2 Kk 0.605 16 Telescoped forms Plate 2 L 0.592 15 Telescoped forms Plate 2 M 0.577 16 Telescoped forms Plate 2 N 0.573 17 Telescoped forms Plate 2 O 0.572 17 0.572 16 210 OTTO. C: GLASER TABLE 3 QUOTIENT EMBRYO SOMITES REMARKS QUOTIENT O.8Lh, —al AL 9 Asymmetrical Too low 0.790 U 13 Asymmetrical Too high 0.789 V 10 Overdifferentiated Too low 0.787 W 13 Asymmetrical Too high 0.768 x 13 Asymmetrical Too high 0.541 Bye 14 Overdifferentiated Too low 0.522 Z 27 Overdifferentiated fulfilled. The relations postulated in advance between the neurocephalic quotients, on the one hand, and aberrancies, de- grees of normal differentiation, and somitic increase on the other, are capable of being verified. The relation which at this time I wish to especially emphasize, is the association of a falling quotient with the multiplication of somites. The quotient correctly understood is a measure of longitu- dinal compression. The somites are recognized as a measure of development. It follows, therefore, that the movements of the quotient are related in time and sense to the serial differentiation to the nervous system precisely in the manner in which they should be reiated, if compression in the longitudinal axis is a condition upon which the vesiculation of the nervous system TABLE 4 SOMITES CASES | QUOTIENT 2 2 1.579 3 2 1.511 4 2 1.163 5 3 1.025 6 3 0.956 7 2 0.912 8 2 0.914 9 5 0.873 10 11 0.845 1 4 0.801 13 2 0.701 15 2 0.621 16 3 0.585 17 2 0.573 DIFFERENTIATION IN THE NERVOUS SYSTEM td depends. The assumption of a ‘causal’ connection between compression and the formation of the brain vesicles is therefore warranted. The justice of this assumption appears still greater when we average the results for each stage of development dealt with in table 2. This was done and the outcome is given in table 4. If now, with the aid of table 4 we construct a curve in which quotients are plotted along the ordinate and the number of somites along the abscissa, the relation between a falling quo- Quotient co Fig. 8 Showing the relation between a falling neuro-cephalic quotient and somitic increase. The curve is constructed by joining the loci of the aver- ages given in table 4. The dots indicate the loci of the individual quotients given in table 2. THE ANATOMICAL RECORD, VOL. 12, No. 2 212 OTTO C. GLASER tient and somitic increase exhibits itself in the most striking manner. On the basis of this curve, figure 8, I infer that during the period of development considered, a rising state of longitudinal compression is one of the conditions determining the differen- tiation of a series of vesicles in the brain. SUMMARY 1. Cranial flexure, although capable of explaining the lateral and ventral differentiations of the prospective second brain vesicle, is nevertheless not related to the general process of vesiculation in the manner in which it should be if the formation of vesicles were dependent upon flexure, as His maintained. 2. According to His, flexure depends on differential growth. 3. According to the results presented in the present paper, vesiculation also depends upon differential growth and precedes flexure. 4. Differential growth, to play a réle in this connection, must be longitudinally effective. Such effectiveness is undemonstrable by the method of His. 5. Effectiveness in the long axis can be demonstrated by com- paring the length of the embryonic head with that of the nervous system. The relation between these two measurements, within the arbitrary limits set by the cephalic rectangle, has been expressed in the form of a fraction. 6. This fraction, the Neuro-Cephalic Quotient, is derived by dividing half the perimeter of the nervous system into the head- length. It tells how many units of head-length are available for every unit of length in the nervous system. 7. The quotient is largest in the earliest stages, and decreases with the progress of development. It is inversely proportional to the number of somites, and, so far as can be determined in the absence of accurate modes of expression, with the degrees of differentiation exhibited by the nervous system. Telescoped forms, and those abnormally over-differentiated, have expectedly low quotients. DIFFERENTIATION IN THE NERVOUS SYSTEM 213 8. Despite the variability which inheres in developmental processes by their very nature, the relations between quotient on the one hand, and somites, or differentiation of the nervous system, on the other, are such as to warrant the conclusion that a rising state of compression in the longitudinal axis is one of the important conditions under which the vesiculation of the embryonic brain takes place. PLATE 1 EXPLANATION OF FIGURES Outlines of heads of diagrammatic 2 to 11 somite chick embryos referred to in table 1. The posterior line through the nervous system is the somitic base line. In embryo A the nervous system did not extend backward as far as the somitic tangent. In this case two base levels are indicated, one used to measure head-length, the other to determine the neural perimeter. 214 DIFFERENTIATION IN THE NERVOUS SYSTEM OTTO C. GLASER avi 215 PLATE 1 ul 0} Poltos oY ‘sqyuotjonb MoO] puBw So}TWOS 7 Z a1qGB4 T 04 &T YQIM sultoj podoose]a], saunapDiIa aO NOILVNVTdXil 6 ULV Id o ULW Id SS “— —= UASVID “9 OLLO WHLSAS SQOAWUN WHHL NI NOILLVILNGYHAAT “ATVI -jyoururAs J[as}t oynqiaysip ABUL ssvUI [BINoU oY} JBY} Jop1o Ul pozooIIp Aj1odoid aq I9AODLOUL PUG OpNyUSvUL UTE}100 B YOBos FsSnUl uoIsso1du10d yey} JSodsns pue ‘gadv oAljo0dser Ioy} 10} YSIY 00} sjyuotyonb oavy “KX pus ‘AA “f) soA1q Ung ‘7, OkAQ UIA BUIPAVSII OpBUT 9q UKO JUIUTE}BIS OU ILO} -o10Y} {pouluLlojJap Used JOU SBY SodvyS o}IWOS JZ oY} LOJ [VUILOU quetgonb ayy, ‘goS3B dATZ00dSed ITO} OJ MO] 00} SjUdTJONbD oAvY X pus ‘A *], “E PUB Z SeTqeF UT UMOYS SY “Ppo}BIJUILOYIp-IoAO oiv 7 puw ‘x ‘A \P SOAIqUIGY “SUILOJ JUBIIOGW SUYADIa AO NOILVNVId Xt £ ALV Id 218 219 § TLWId MASVID *O OLLO WAULSAS SNOANMAN AHL NI NOILVILINGAY Adda THE CHOROID PLEXUS WITH SPECIAL REFERENCE TO INTERSTITIAL GRANULAR CELLS JOHN SUNDWALL Department of Anatomy, University of Kansas TEN FIGURES During a comparative study of the structure of the choroid plexus in various mammals, in association with Dr. Harvey Cushing at the Hunterian Laboratory, Johns Hopkins Univer- sity, I observed the presence of very prominent interstitial granular cells in the choroid plexus of the ox. They are especially numerous in the ox and are present to a less degree in the sheep and swine. I have failed to find any reference to these cells in the literature on the choroid plexus, and it is primarily to report them that this paper is submitted. I have included, as well, my observations on the structure of the epithelial layer of cells, in view of their now generally accepted connection with the secretion of cerebro-spinal fluid. GENERAL ARCHITECTURE The choroid plexus of the ox is similar in structure to that of other mammals, as generally described. Tufts of blood ves- sels varying in size and thickness of the connective tissue coats are seen covered by a single layer of cuboidal cells. The various elements making up the choroid plexus are clearly differentiated in tissues fixed in formalin-bichromate solution and stained in Van Gieson’s. The yellowish-brown stained surface cuboidal cells are sharply demareated from the red stained con- nective tissue which surrounds the blood vessels. In some tufts the connective tissue coats which form the walls of the blood vessels measure 0.2 mm. in thickness, while in others they form By | Dae, JOHN SUNDWALL only a thin membrane between the vessel lumina and the surface cuboidal cells. Much variation exists also in the size of the lumina (fig. 1). Generally speaking, the vessels of the choroid plexus may be divided into two groups: 1) The vessels with the thick walls possess as a rule narrow lumina and have the same structure in general as do small arteries. Weigert’s stain shows a well de- veloped tunica elastica interna, which in most instances is cor- Fig. 1 Choroid plexus of the ox. a, artery; 6, vein or sinus. Microphoto- graph, oc. 10, obj. 16 mm., B. & L. Tech: Bensley’s aleohol bichloride bichro- mate solution, haematoxylin and eosin. rugated. Fine, waving elastic fibers may be seen in the tunica media, which is well developed. The fibers may be further traced to the cuboidal cells and are frequently seen to form a basement membrane for these cells. 2) The second group of vessels are those with comparatively thin walls and wide lumina. These may be regarded as veins or sinuses. ‘The walls are com- posed almost entirely of white fibrous connective tissue and cannot be differentiated into intima, media, and adventia, but are similar in structure throughout. Only here and there are CHOROID PLEXUS 2a seen fine elastic fibers. No muscular walls can be made out in this second group. The walls contain numerous small vessels and capillaries (Vasa vasorum) of varying calibres. They are rich also in nuclear elements, such as endothelial cells, connective tissue cells and interstitial granule cells (fig. 1). A discussion of these interstitial granule cells follows that of the cuboidal surface cells. EPITHELIAL CELLS The epithelium forms a single layer of cuboidal cells. This observation agrees with those of Meek (’07) and others who de- seribe a single layer of cells. Luschka (’55), Findlay (’89) and Kolliker (96), however, state that several layers are present. The cells on some tufts are more or less flattened while on other tufts they are found to be elongated. This condition depends upon the degree of distention of the vessels. Generally the epi- thelial layer presents on the ventricular surface an even, unin- terrupted surface, so closely and evenly are the cells arranged. Tufts are present, however, where slight indentations projecting basalward between the cells are seen. Thus the rounded free ends of the cells, in cross sections, give the epithelial layer a corrugated appearance. Frequently cells are seen with bipar- tate free ends. In the ordinary fixations and stains the cytoplasm in the ma- jority of the epithelial cells stains evenly and compactly through- out. A very finely granular substance is distributed throughout this cytoplasm. No intracellular net work is seen. Vacuoles are frequently present. The nuclei vary somewhat in form. As a rule, they are spherical. In the more or less flattened epi- thelium they are oval in outline, with their long axes parallel to the surface of the tufts. Much chromatin is present in the nucleus although it does not stain solidly throughout, as is the case in many gland cells. Several large chromatin masses, as a rule, are seen in each nucleus surrounded by numerous finer granules. One or two nucleoli may be seen. These, however, are difficult to differentiate in the ordinary stains from the 224 JOHN SUNDWALL larger chromatin masses. The nuclei occupy the central portion of the cells, perhaps somewhat more basalward than towards the free surface. Examination of many choroid plexuses does not reveal any marked variation to this position. I did not observe a nucleus compressed against the basal end of the cell, as is seen in many gland cells. For a more detailed study of the surface epithelium, choroid tissues were fixed in Bensley’s aleohol bichloride bichromate solution and formalin bichromate solution. Sections were stained with iron haematoxylin, copper chromehaematoxylin, Bensley’s (11) neutral gentian and safranin acid violet. In all these stains the epithelium stands out prominently. The cell membrane on the surface is seen as a relatively thick line, at times it appears as a double contoured line. No striations are seen in this cell membrane. After some fixations, where there is evidence that the process has not been perfect, the cell membrane appears as a thick margin, still retaining the original shape of the cell while the cytoplasm is shrunken away from it. In such preparations, the impression is given that these cells have thick, more or less unyielding membranes. No cement substance is present between the cells. Secretion granules, such as are seen in the pancreas, parotid, lachrymal, gastric and other glands, are not seen in the epithelial cells of the choroid plexus. How- ever, after staining with neutral gentian or safranin acid vio- let, one finds cells which possess a few deeply stained granules that simulate in structure and staining characteristics the secre- tion granules of the above named gland cells. Such cells are not numerous, and the granules are only sparsely present, not more than one-half dozen have been counted in each cell. The nature of these granules has not been determined. They may represent nuclear substance within the cytoplasm. Occasionally one observes as well, large bodies near the nucleus which take the nuclear stain deeply. These may be interpreted as para- nuclei and have been observed in various cells. The epithelial cells possess numerous vacuoles and canaliculi. Others have observed vacuoles in the epithelial cells of the choroid plexus. Meek found fat globules in these cells in the rabbit, CHOROID PLEXUS 225 which in prepared sections, owing to the dissolving action of the various preparation fluids, appeared as vacuoles. He proved that these were due to the former presence of fat, by staining with Sudan III choroid cells which had not been subjected to the dissolving action of alcohol and xylol. Globules, or vacuoles, have been observed in this epithelium by Luschka, Findlay, Studnicka, and Galeotti, which, according to these observers, are evidences of the vesicular type of secretion. On the other hand, Pettit and Girard are prone to regard the vacuoles as abnormal structures due to mechanical injuries or post mortem changes. . Numerous intracellular spaces were seen in my preparations notwithstanding that the greatest care was exercised to avoid mechanical injury to the cells. The tissues were removed and fixed almost instantly after the animal had been slaughtered, so that no postmortem changes could have occurred. These spaces are in the form of globules or vacuoles and canaliculi. The vacuoles may be seen in any part of the cell, either at the base or the free margin. They may be round or oval. Occa- sionally a row of these oval vacuoles are seen at the base of the cell. The intracellular canalicular apparatus of Holmgren ('02), Bensley (’10) and others, may be seen in the cytoplasm as branch- ing canals which frequently can be traced to the nucleus and partially encompassing it. They are entirely intracellular, as the branches terminate within the cytoplasm (fig. 3a). That the finer vacuoles are cross sections of these canals is probable. Whether they bear any relation to the secretory ac- tivity of the gland, I am unable to say at the present time. That the larger vacuoles do not represent spaces remaining from dissolved fat globules is certain, for I have been unable to dem- onstrate the presence of such fat globules in the epithelium of the choroid plexus of the ox (figs. 3, 6, ¢). The description so far given for the epithelial cells is confined to those which are generally observed. Studies of numerous choroid plexuses, however, show various types of cells. In ad- dition to those already described, one sees in fixed preparations 226 JOHN SUNDWALL cells whose cytoplasmic area is composed almost entirely of vacuoles. The cytoplasm is manifested only between the vacuoles, forming a network surrounding the spaces. In another type, the entire cell is seen to be enlarged. It may be two or three times as large as the general type. It bulges out and is rounded as a consequence of its contents (fig. 2). This type shows that a confluence of vacuoles has taken place. This cell stains faintly because of the relative decrease in the cytoplasm. Fig. 2 Choroid plexus of the ox. Two types of epithelial cells are seen in the section: large, faintly staining, distended cells; and small deeply staining cells, which is the type usually making up the epithelium. Microphotograph, oc. 10, obj. 4mm., B. & L. Tech.: Same as 1. Only at the base of the cell is seen a narrow ragged zone of com- pact cytoplasm. In these large vacuolated cells, no changes are observed, as a rule, in the form and position of the nuclei, which are generally centrally located; however, I have observed the nuclei in the apices of many of them. The large bulging vacuolated cells may be seen between two smaller cells whose cytoplasm is compact. Again they may constitute the majority of cells on a particular tuft or villus. CHOROID PLEXUS Bar That these various cell types belong to the same category and that they represent different secretion stages of the same cell type is evident. From the study of these cells in fixed sections, one is con- strained to believe that the secretion of the epithelial cells of the choroid plexus is a vesicular one. The secretion substance makes its appearance at the base of the cell as minute vesicles. Later they are seen throughout the cytoplasm. A confluence of these vesicles takes place to form large vesicles. At the same time the cell enlarges, becomes rounded and bulges out between the resting cells. Then the secretion passes out into the ven- tricles without a break in the continuity of the cell membrane. Fig. 3 Three cells, selected, showing canals and vacuoles within the cyto- plasm. a, intracellular canaliculi; 6 and c, vacuoles. Drawing somewhat diagrammatic, oc. 10, obj. 1.9 mm., oil immersion, B. & L. Tech.: Bensley’s alcohol bichloride bichromate solution, neutral gentian. That a continuous secretion is taking place is apparent from the fact that all the various secretory stages of these cells may be seen in one choroid plexus. One finds, however, some tissues in which the vacuolated cells are much more promi- nently present than they are in others. As a rule, the epithe- lium is of the general non-vacuolated cuboidal type which is described first. The processes of secretion in the epithelial cells are in no way comparable to those of the duct gland cells,—salivary, pan- creas, lachrymal glands. Secretion granules, the antecedents of these granules, and nuclear changes as seen in other gland cells are not demonstrable in the epithelium of the choroid plexus. The secretion substance of the latter makes its appearance as minute droplets within the cytoplasm and no antecedent secre- tion substances have been observed. A somewhat similar THE ANATOMICAL RECORD, VOL. 12, No. 2 228 JOHN SUNDWALL method of secretion has been observed in the kidney by Gur- witsch and others. In other respects the secretion methods of the two organs,—choroid and kidney—are similar. In both large amounts of fluid are secreted (excreted) with but relatively little changes in the cells concerned in the secretion. The latter differs from the former in that no foreign substances, such as injected indigocarmin, potassium ferrocyanide, iron-ammonium citrate, prussian blue, have been observed to pass through the choroid epithelium into the ventricles. Experiments were per- formed on dogs by Prof. 8S. A. Matthews and myself in order to find some substance that, when injected into the blood, could be detected microchemically in the epithelium of the choroid. Our results will be published later. The action of the choroid cells may be compared also, to that of the endothelial cells of blood vessels in the formation of lymph, providing we accept the theory that physical processes alone— filtration, diffusion and osmosis, do not explain all the phenome- non of lymph secretion, but that the endothelial cells are con- cerned in the secretion of lymph, in which case a large amount of fluid is produced without any distinct observable histological changes on the part of these endothelial cells engaged in lymph secretion. Meek found that with an increase of cerebrospinal fluid fol- lowing muscarin injection, the epithelial cells had increased in height and certain clear spaces formed in the apical ends of the cells. His observations corroborated in a way the observations of Pettit and Girard (’02). [ have not confirmed, in the opossum, the observations of Meek in this respect, but my observations are limited to a few adult animals. One must presume that the choroid plexus has an autonomic innervation like the salivary glands, ete., in or- der to secure this phenomenon. I have not demonstrated to my own satisfaction, either by the silver method or by vital methylene blue staining, that the cells of the choroid plexus are innervated. [ am inclined to hold that the choroid plexus cells and their activity in the secretion of cerebro-spinal fluid belong to the CHOROID PLEXUS 229 category of endothelial cells and cannot be compared to duct gland cells derived from epithelium. Definite intercellular spaces are also observed. These may be in the form of mere indentations between the surface ends of the cells or a definite cleavage may be seen reaching to the basal end of the cell, where it frequently appears continuous with other canal-like structures which enter the deeper connective tissue walls (fig.4). Occasionally the intercellular sp*ces are globular in outline. These observations do not agree with those of Meek and others, who hold that the epithelial cells are so closely ap- pressed that intercellular spaces do not normally occur and that Fig. 4 Composite drawing of cells showing: a, intercellular canals; } and c, sub-cellular and interstitial canals; d, blood vessel. Drawing somewhat dia- grammatic, oc. 10, obj. 1.9 mm., oil immersion, B. & L. Tech.: Same as 3. when they are seen, they are due to faulty technique. Stud- nicka (’00), on the other hand, holds that these spaces are seen in the choroid plexus of the shark. While I have not observed these intercellular and sub-cellular canals in living tissue, I am not convinced that they are always to be regarded as artifacts. The frequency with which they are seen, the variations observed in their outlines, point to the fact that they are normally present. The canals are frequently seen te continue as such in the connective tissue walls and apparently communicate with lymphatics. To maintain that they are nor- mally present does not seem inconsistent when one compares the choroid plexus with other serous endothelial cells. In my opinion, these spaces can be readily compared to the stomata 230 JOHN SUNDWALL described by v. Reklinghausen, Klein, Dogiel, and others be- tween the endothelial cells of the peritoneum and by Ludwig and Dybkowsky in the pleura. The silver precipitation methods were utilized in the study of these intercellular canals. A heavy black precipitation filling the entire epithelial cell occurs when the tissue is fixed in Kopsch’s fluid. Thus when viewed through the low power of the micro- scope, this deposit in all the surface cells forms a black border around the choroid tufts. Projecting from the basal margin of this black border (the epithelial cells) into the connective tissue are seen numerous canal-like deposits which suggest the inter- cellular and subcellular canals. However, further observation is essential before I can come to any definite conclusion in this particular. The very heavy deposit of silver within the cells makes the observations difficult and one is not always sure as to whether he is dealing with definite tissue structures or with artefacts. Further investigation is essential before one can state just what the function of these intercellular canals is. When com- pared again with the generally accepted function of peritoneal stomata, it is suggested that these canals may be absorptive in character, that is, the cerebro-spinal fluid may pass back into the circulation through them. If such is true, the choroid plexus then may be regarded as both a secretory organ and an absorp- tion organ. On the other hand, the presence of the canals sug- gests another possibility, and that is, that they may function directly in the production of cerebro-spinal fluid. Should we accept the mechanical theory of secretion, it would not be fan- tastical to suppose that much of the fluid may be regarded as a transudate passing directly from the connective tissue surround- ing the lymphatics and blood capillaries through these small sub- and inter-cellular canals directly into the ventricles, with- out passing through the epithelial cells. According to this hy- pothesis, the epithelial cells of the choroid plexus alone may not be concerned in the production of cerebro-spinal fluid. In view of the fact that large amounts of cerebro-spinal fluid may be secreted in a short time (200 to 300 ee. and possibly more in 24 CHOROID PLEXUS 231 hours in the human, Cushing 714), one might naturally expect to see more histological evidence of secretory activity on the part of the epithelial cells, providing these cells are solely concerned in the production of the fluid. Another observation that sug- gests this second hypothesis is that in the choroid plexus of the ox, numerous mast cells apparently pass directly from the blood vessels through the connective tissue wall and between the epithelial cells into the ventricles. Against this view that the cerebro-spinal fluid may be regarded in part as a transudate is the fact that it differs from lymph, Fig. 5 Mitochondria in the epithelial cells. Drawing, oc. 10, obj. 1.9 mm., oil immersion, projection apparatus. Tech.: Bensley’s acetic osmic bichromate Solution, anilin fuchsin methyl green. plasma, tissue juices and serous cavity fluids in the relative amounts of protein substance. Cerebro-spinal fluid is almost free from protein, while the other fluids contain much more. That the choroid plexus is responsible for the formation of cerebro-spinal fluid was suggested by Faivre a long time ago. Luschka, a little later, agreed with him, and practically all ob- servers since, including Pettit and Girard, Studnicka, Galeotti (97), Carazzani, Meek, Mott (710), Goldmann (713) and Cush- ing, regard this structure as fundamental in the secretion of cerebro-spinal fluid. Cushing saw drops of the fluid exuding from this tissue. Goldman noted the extrusion of glycogen in the form of globules from the cells. Weed, in connection with others, suggests a dual origin of cerebro-spinal fluid—from the choroid plexus and from the perivascular systems of the nervous tissue. Bon JOHN SUNDWALL Certainly much more work must still be done in order to solve the method of secretion which takes place in the choroid plexus. When the tissue is prepared by Bensley’s acetic osmic bichro- mate, anilin fuchsin, Methyl green method, the epithelial cells show numerous mitochondria. These are in the form of very short bacillus-like rods and tend in a way to arrange themselves in irregular rows, reaching from the base of the cell to the sum- mit. Of course these rows are by no means so definite and clean cut as I (16) have observed in the duct cells of the lachry- mal glands (fig. 5). In the vacuolated cells, the mitochondria are seen in the cytoplasmic reticulum between the vacuoles. INTERSTITIAL GRANULAR CELLS This type of cell is characterized by the presence of numerous large granules which completely fill the cell. The nucleus is as a rule completely obscured by these granules which simulate in form and staining characteristics the secretion granules of gland cells. These granular interstitial cells were found to be present with- out exception in the choroid plexus of the ox, but varied in num- ber with different animals. In some, they were especially num- erous, while in others only a few were found. Between these two extremes there was every gradation. In the sheep and swine they are not so numerous. In the choroid plexus of other ani- mals examined—rabbit, guinea pig, dog, and human—these cells have not been observed. The following description of the interstitial granule cells will be confined to those observed in the choroid plexus of the ox. The cells vary in size and shape. The average size is 20 micra. The majority are more or less spherical or oval in outline. Many, however, are angular, others are elongated or irregular and pos- sess processes extending from the cells. They are situated in the connective tissue walls of the vascular tufts between the vessels’ lumina and the surface epithelium. In some fields (low power microscope), as many as 30 of these cells have been counted (figs. 6, 7, and 8). CHOROID PLEXUS 233 The thickest connective tissue walls, those surrounding the arteries, contain the largest number of these cells, which are found at varying distances from the lumina of the vessels. Some are seen in close contact with the endothelial cells of the vessels and in some instances, processes of the granular intersti- Fig. 6 Stroma of choroid plexus of ox. ‘The numerous interstitial granular cells—mast cells—are deeply stained. Microphotograph, oc. 10, obj. 4 mm. B.& L. Tech.: Same as 3. tial cells are seen to project between two endothelial cells into the lumen. The other extreme is seen where processes of these cells lying directly under the epithelial layer of the choroid plexus project upward between two epithelial cells into the ventricle. Between these two extremes the granule cells are seen in the con- nective tissue at different levels. None of these cells have been observed lying free either within the vessel lumina or on the ventricular surface of the choroid plexus. 234 JOHN SUNDWALL Fig. 7 Interstitial granular cells—mast cells—in the connective tissue wall. a, Surface epithelium; b, The deeply stained interstitial granular cells—mast cells—are seen at varying distances from the lumina of vessels. Some are directly underneath the epithelium with processes projecting between the epithelial cells. Others are seen directly underneath the endothelium of vessels; c, artery. Drawing, oc. 10, obj. 16mm., B. & L. projection apparatus. Tech.: Bensley’s alcohol bichloride bichromate solution, Unna’s polychrome methylene blue. AY oI ° Fig. 8 Four selected interstitial granular cells—mast cells—showing varia- tion inform. Drawing, oc. 10, obj. 16mm., B. & L. projection apparatus. Teeh.: Same as 7. CHOROID PLEXUS Zan The cells are readily seen in both fresh tissue mounted in serum or salt solution and in fixed preparations. Bensley’s so- lution—equal parts of (a) saturated solution of mercuric chloride in 95 per cent alcohol and (b) 2.5 per cent aqueous solution of potassium bichromate—proved to be the most satisfactory fixing solution for the granular cells. They are fairly well fixed in alco- hol. Fluids containing much acid destroy the granules, conse- quently Zenker’s solution cannot be used. In Bensley’s acetic osmic bichromate solution, the granules are only partially con- served. This fact, however, proves of great value because then the other constituents of the cell can be made out,—nucleus and intergranular substance. The stains first utilized for the study of these cells were Bens- ley’s neutral gentian and neutral safranin. The granules in these dyes stain deeply and hold on to the stain with much tenacity. Even after the section as a whole has been differentiated in aleo- hol-clove oil to the extent that practically all the stain has been removed, the granule cells remain deeply stained. And so num- erous are these granules within the cell, as a rule, that the entire cell appears as one deeply stained mass. It is only after ex- tended differentiation or in the processes of the elongated cells that the individual granules can be made out. In these stains the nuclei in most cells are completely obscured by the deeply stained granules. Only in those cells where the knife has passed through the nucleus or in the smaller cells where only few granules are seen around the nucleus is the latter seen. It. occupies a central position. It is oval in outline and vesicu- lar,—thus staining faintly. Only a small amount of chromatin is present which is distributed throughout the nucleus appearing as fine irregular clumps. No nucleolus is observed. When these nuclei are seen through the low power of the microscope they appear as transparent or faintly opaque areas in the center of the cells surrounded by the deeply stained granules (fig. 9, a). The presence of so many of these large round or oval mono- nuclear cells possessing numerous granules which simulate, both in size and staining characteristics, secretion granules suggested 236 JOHN SUNDWALL at first the possibility of the choroid plexus containing some type of endocrinous gland (fig. 6). Extracts of the choroid plexus were made in the usual manner and injected intravenously into a dog. Outside of the usual depressor reaction,—a frequent phenomenon accompanying in- jections of animal tissues,—no specific or particular reaction was observed. Dixon and Halliburton (13) on the other hand claim that choroid extract when injected intravenously increase mark- edly the flow of cerebro-spinal fluid. Our observations, how- Fig. 9 a, Interstitial granular cell, in which the large basophilic granules are not conserved. The mono-nuclear character of the cell is clearly seen. Note the fine intergranular network of cytoplasm. b, Broken mast cells, the granules of which are fuchsinophilic. Drawings, oe. 10, obj. 1.9 mm., oil immer- sion, B. & L. Tech.: Same as 5. ever, were confined only to one experiment and consequently will not warrant any conclusion on this phase of the subject. The probability that these interstitial granular cells are mast cells early suggested itself, especially after other stains had been used. However, I have not seen them so constantly and prominently present in any other tissue. Their presence in such large numbers may suggest that they have some function other than that generally ascribed to mast cells. However, our knowl- edge of the nature and function of mast cells is very superficial. Numerous theories have been advanced regarding the réle they play in animal life. Many hold that they are in a way unicellu- lar glands concerned in some type of secretion. If such be the CHOROID PLEXUS Dab case, one may still regard these interstitial granular cells of the choroid plexus as having some unknown secretory function. At any rate, it was later determined that they possess the same mor- phological and staining characteristics as the so-called ‘his- tiogenen Mastzellen’ of Ehrlich and others. In view of these facts, the interstitial granular cells of the choroid may be re- garded, for the present, at least, as belonging to this category, although their particular function here is unknown. Since Waldeyer (75) first described a particular group of con- nective tissue cells possessing granules, to which the term ‘plasma cells’ was applied, and later Ehrlich (’77) and his assistant West- phal (’80) recognized a still more limited group whose large granules showed a strong affinity for basic dyes and to which the term ‘mast cells’ was applied, much interest has been shown in these interstitial granular cells. In the literature on this subject, mast cells are generally de- scribed as interstitial cells more or less round in contour, the cytoplasm of which is made up of large granules which have a marked affinity tor basic stains, Mallory (14), Schafer (12), Rauber Kopsch (’06). Most authors fail to differentiate be- tween the two types first recognized by Ehrlich and Westphal, who referred to them as ‘mastleucocyten’ and ‘Histogenen mast- zellen.’ Some writers describe the former and some the latter, when a definition of mast cells is made. Regarding the two types, Prenant, Bouin, Maillard (’04) state: “Les cellules & granulations basophiles du sang sont les mémes que les cellules nutritives que nous retrouverons dans les tissu conjonctif sous le nom de Mastzellen ou cellules-engrais.’’ The two cell types were recognized by Pappenheim (01) who dis- cusses the origin of each, and by Maximow (’06) who states that the relation of the two types is not clear. The distribution of mast cells, according to many investiga- tors, is wide. This is true not only of the distribution in the individual, but also throughout the various orders of animal life. They have been described both in invertebrates, such as the cellules mucoides, described by Guenot in ‘Gastropodes Pulmones’ and in every order of vertebrates. It was the opin- 238 JOHN SUNDWALL ion of early investigators that mast cells were found in the latter only in batrachians, where in the Urodeles they were seen in enormous numbers. Now it is generally conceded that they are found in all vertebrates. They have been described in triton, frog, turtle, rabbit, calf, man, and in fact, all species of vertebrates. In the individual body they are distributed as follows: In the blood-basophilic leucocytes; connective tissue mast cells have been described in the connective tissue layers of the skin and mucous membrane. They are numerous in the tongue, in the septa of ‘arious glands, in the lungs. Arnold (714) found that mast cells are greatly increased in numbers in the frog’s tongue after in- duced passive congestion. Mincheimer (’95) saw them in the testes of horse, rat and pig. He failed to note them in these organs of the deer, sheep, dog and rabbit. Korybutt Daskie- wiez (’78) describes them along the nerve fibers in the frog. MeKibben (14) states that they may readily be mistaken for nerve cells as observed in the nasal region and meninges of nec- turus. They are found in bone marrow, adipose tissue, along blood vessels, in newly formed connective tissue. These mast cells make their appearance early in embryonic life. They have been observed in the 9-day chick and in early calf embryos. These cells are much more clearly seen, as a rule, in the embryo than in later life. I have failed to observe in the literature on this subject refer- ence to these large, prominent interstitial granular cells—mast cells—in the choroid plexus of the larger mammals. Haeckel (59) refers to certain interstitial cells in the choroid plexus of embryonal mouse and dog, which, according to him are similar to those seen by Schultze (’56) in the gelatinous connective tissue of tunicates and medusae. Haeckel considers these wandering cells and, according to him, they are filled with minute globules of fat. [ have stained the choroid plexus of the ox with a view of determining the presence of fat in the mast cells. Sudan III, Scharlach Rot—in aleohol or as Herxheimer’s stain, and the Nile blues were used. So far, I have failed to demonstrate the CHOROID PLEXUS 239 constant or prominent presence of fat in these mast cells. Only rarely are fat droplets seen. Hence the mast cells in the choroid plexus of the ox differ from those cells described by Haeckel. They are also different from the other mast cells of the con- nective tissue type in which fat and lipoid substance have been found, as claimed by Pappenheim, Posner, Lombardo and others. Huguenin (712) saw numerous lipoid granules in mast cells in a case of status lymphaticus. According to Ciaccio (13), lipoid and fat are found in these cells only under pathological conditions. Flemming (’71, ’76, ’79) and Hammar (’95) were of the opinion that fat was not deposited in mast cells. Cer- tainly fat is not demonstrable as a constant constituent of the mast cells of the choroid plexus. Reaction to mucin stains. One of the first stains used in the study of these cells was muchaematein, in which the granules stained a deep blue. No other elements in the tissue were stained. The presence of so many round or oval cells specifically stained in muchaematein or mucicarmin was another factor that suggested a glandular secretion phase to these interstitial cells. Of course, the presence of mucin naturally suggested itself. I (16) have shown, however, that secretion granules—lachrymal gland—do stain specifically in these mucin stains, although we have no evidence that mucin is secreted by the lachrymal gland. Others have observed that the granules in mast cells are stained with mucous stain. Raudnitz (’83) held that because of this reaction, the granules represent a stage in mucinous degeneration of the ceil. Hoyer (90) held that the granules were of a mucin- ous nature. Schaffer concludes that as the mast cells take the seme stain as do cartilage cells, the former contain a chondroitin sulphuric acid substance. On the other hand, Pappenheim, Schwenter, Trachsler and others held that the granules of mast cells have no relation to mucin. Ehrlich observed that the granules of mast cells are much more resistant to water than are mucous granules. An interesting phenomenon in the study of muchaematein stained mast cells is the variability in the intensity of the stain- ing reaction of the granules. Some cells take the stain much 240 JOHN SUNDWALL more deeply than do others. One frequently secures choroid plexus in which the cells as a whole stain much more quickly and deeply than do the similar cells in other choroid tissue. This variety in intensity of staining with muchaematein may repre- sent various stages in the formation of granules. No other ele- ments in the cell are stained. Preparations according to this method are of great value in the study of the individual gran- ules. These are seen to fill the entire cell and where processes are present, the granules extend into them to the very ends of the processes. They vary in size. Reaction to basic stains. When stained in methylene blue, toludin blue, methyl green, etc., the granules are deeply stained. They hold these stains with such tenacity that when the sec- tions are differentiated in aleohol, practically all the stain may be removed from the section before there is any perceptable loss of stain from the granules. The nucleus, when seen at all, occu- pies a central position in the cell and appears as an opaque or semitranslucent structure. Iron haematoxylin and copper haematoxylin stains. The gran- ules do not retain these stains after differentiation. In fact, the mast cells are among the first of the structures to give up the stain during the process of differentiation. The granules of the mast cells differ in this respect from secretion. granules of many serous cells. The latter retain both stains deeply after practi- rally all the iron and copper reactions have been removed from the section as a result of the differentiation. Polychrome methylene blue stain. Here the granules stain metacromatically—a deep violet, while the remainder of the choroid tissue in general is only very faintly stained. The epi- thelial layer is tinged green. Here, as observed in other stains, the granules in the mast cells are so densely stained that the entire cell appears as a dark violet mass simulating an artefact. Pro- longed differentiation in alcohol reveals the characteristics of the individual granules, especially in those cells where the granules are not so numerous. In many cells a translucent central area is observed which marks the position of the unstained nucleus. CHOROID PLEXUS 241 In this fixation (Bensley’s) and stain, the nucleus is never defi- nitely seen because of the numerous granules that obscure it. Alcohol fixation. The granules are not so well conserved in 70 per cent alcohol, although according to many observers, alco- hol is one of the best fixatives for mast cells. This method of fixation, however, has its value, as the nuclei of the cells are plainly seen, owing to the fact that the granules are not well pre- served. The nucleus is stained green (Polychrome methylene blue). It is very vesicular and is as a rule oval in outline. No nucleolus is present, as was also observed by Dantchakoff (98), who states that no nucleolus is present in the nucleus of mast cells. Variable sized chromatin granules are present. As a rule one to four or five large chromatin masses, one to two micra in diameter, are seen in each cell. Interspersed among these chromatin granules is the fine chromatin dust. The chromatin is only sparsely present, which accounts for the vesicular appear- ance of the nucleus. The granules of the mast cells are almost totally destroyed when fixed in Bensley’s acetic osmic bichro- mate solution. Here the nucleus can be readily observed. I was unable to demonstrate the presence of mitochondria in these affected cells after fixing in the above solution and staining with anilin acid fuchsin, methyl green. Occasionally, however, mast cell granules were seen which, instead of being stained green by the methyl green, were stained deeply red by the fuchsin (fig. 9, b). When the tissue is prepared according to the method utilized by Cowdry (716) and Seott (16) for study of mitochondria, the granules of the mast cells are well preserved and are stained deeply green in contrast to the red stained mitochondria so abundantly present in the surface epithelium. I did not ob- serve mitochondria within the mast cells. Had any of these minute fuchsinophilic granules been present, they should have been observed readily in contrast to the deep green stained mast cell granules. This observation is corroborated later on in the vital stains. 242 JOHN SUNDWALL VITAL STAINING Choroid tissue was obtained by gently separating the plexus from its attachment in the ventricle immediately after the ani- mal was slaughtered. Small bits of the choroid were then placed in the following solutions: Neutral red, one part in 15,000 of isotonic salt solution; new methylene blue, one part in 10,000 of isotonic salt solution; janus green, Metz, 1 to 15,000; poly- chrome methylene blue, Unna, | to 15,000; and pyronin, 1 to 1,000. Small pieces were removed from each of these solutions, mounted in the respective solutions and studied under the microscope. The remaining tissues were fixed in ammonium molybdate solution. The following observations were made: Neutral red stain. The choroid plexus is rapidly stained with this dye. Macroscopically, the entire tissue is stained deeply red. Microscopic examination reveals the following characteris- tics: The surface epithelium is stained deeply red, the stain is confined to the cytoplasm and is diffuse throughout. The numerous, minute, highly refractive granules which were ob- served in fresh, unstained tissue, stand out prominently in the diffusely red stained cytoplasm. The nucleus which is unstained is spherical and occupies a central position in the cell. Numerous mast cells are seen, the granules of which stain rap- idly and deeply red. The large oval nuclei of the mast cells remain unaffected by the stain. As a rule, they are much more readily seen in this preparation than in the permanent prepara- tions. Contrary to the observations of Arnold, I noted some activity on the part of these mast cells when mounted in serum and enclosed within a stage incubator. One cell was seen to completely divide within the period of one and one-half hours. This particular cell, which was oval in outline at first, was seen to elongate itself by processes extending from both ends (fig. 10). The formation of new granules in the processes was concurrent with the development of the processes. With the elongation of the cell, a constriction occurred in its central portion, which con- tinued until the cell was completely divided, and with the divis- ion of the cell, nuclei appeared in both halves. Of course it was CHOROID PLEXUS 243 impossible to observe in this stain the method of nuclear di- vision. Arnold saw no changes in the form and position of mast cells within a period of from twenty-four to thirty-six hours. Kanthack and Hardy held that the ‘Histiogenen’ mast cells are stationary. While Lowenthal, Maximow, Pappenheim and Weidenrich ascribed to them an amoeboid movement. Maximow saw alterations in the form of mast cells in inflam- matory tissue. In numerous other cells, processes were observed to extend out from the cell bodies and to work themselves between the con- OP R $d Fig. 10 Outline of changes observed in a living interstitial granule cell— mast cell—during division. The time required was one hour and fifty-five min- utes. a, 4.385 p.m.;b, 5.05 p.m., c, 5.50 p.m., d, 6.10 p.m., e, 6.30 p.m. Drawing, oc. 1-, obj. 1.9 mm., oil immersion, B. & L. Tech.: Fresh choroid tissue mounted in serum, vital staining with neutral red, electric incubator for microscopic stage, temperature, 98.5°. nective tissue elements. Some of the processes developed within one-half hour. The development of the processes was charac- terized, as a rule, by a continuous out-flowing of granules from the cell body. Granules were seen, however, to develop in the processes some distance from other granules, thus apparently having their origin directly from homogeneous cytoplasm. In attempting to determine the origin of these granules, I have come to no conclusion. In many instances the granules ap- peared to divide. However, as noted, granules seemed to make their appearance directly from homogeneous cytoplasm. Many observers hold that the granules are derived from the THE ANATOMICAL RECORD, VOL. 12, No. 2 244 JOHN SUNDWALL nucleus. Downey states that the chromatic substance comes from the nucleus, through its wall, and then comes in contact with the primary granules of the cytoplasm, after which the basic and metachromatic staining of the granules follow. My observations show that the granules may form in the processes some distance from the nucleus. Just what relation they bear to the nucleus is impossible to state. The fact that they take the nuclear stain is responsible for the assertions that they have a nuclear origin. Polychrome methylene blue. Here the epithelial cells are char- acterized, when viewed from the surface, by relatively thick unstained cell boundaries which mark the polyhedral cell bound- aries. The unstained spherical nuclei occupy the central por- tion of the cells. These are surrounded by unstained cytoplasm with highly unstained refractile granules. The nuclei, however, begin to stain early in this dye, thus indicating the toxic effect of the stain. The granules of the mast cells rapidly stain metachromatically and the nuclei, as in the case of neutral red, are unstained. Later they begin to stain, as in the case of the epithelial cells. Janus green. After the tissue has been in this solution for a few moments the connective tissue elements are stained, under macroscopic observation, metachromatically a pinkish violet or purple, while the epithelial cells are stained blue. When the tissue is studied by means of the oil emersion, it is seen that the mitochrondria of the epithelial cells are deeply stained blue, while the other elements of the cell are unstained. The mito- chondria are numerous and show the same characteristics in both structure and distribution as observed in fixed preparations. A narrow, definite zone of homogeneous, unstained cytoplasm immediately surrounds the nucleus, in which no mitochondria are seen. This frequently gives to the nucleus a double contour appearance. The mitochondria in the epithelial cells of the choroid plexus are indeed numerous. They are found in such great numbers that under low power the cytoplasm appears to be a solid blue throughout. It is only under the oil immersion that the minute CHOROID PLEXUS BAY individual granules are observed. These granules vary in form. Some are spherical, others are slightly elongated, either slender or thick, others again are comma shaped. In no instance are long rods seen. It is the mitochondrial granules that are seen as the minute, highly refractive granules in fresh tissue, as well as in the various other vital stains used, where they remain unstained. In the janus green solution all the granular elements of the epithe- lial cells are stained blue. It is my opinion that the granules that have been described by previous investigators as secretion granules are in reality mitochrondria. The mast cells are unaffected in this dye. Only rarely does one see granules stained with Janus green. Frequently one ob- servers a fine, intergranular deposit of blue stain between the granules of the mast cells. I was unable, however, to make out definite mitochrondria in this deposit. Neutral red and Janus green. When both neutral red and janus green, mixed, are applied to fresh choroid tissue a very interesting picture results. The epithelial cells are deeply stained blue owing to the numerous mitochondria they possess. On the other hand, the mast cells are stained deeply red as a consequence of the affinity between the granules and neutral red. Careful examination of numerous mast cells did not re- veal the presence of blue stained intergranular mitochondria. In fact, the complete absence of the latter was the striking and surprising feature, in view of the claims of Arnold and Dubreul that mitochondria are found in mast cells. Tochaschin (’12) reports staining of true mitochondrial elements in fibroblasts and clasmatocytes by isamine blue and trypan blue. In this preparation, when enclosed within a warming stage, I observed processes projecting themselves outwards from cells in which granules concurrently appeared but no mitochondria were ob- served. This observation is against the view that some have held—namely, that the granules of mast cells have their origin from mitochondria. These observations confirm those already described for the fixed preparations. Methylene blue. Two preparations of this stain were used. Methylene blue rect. Griiber’s, and Methylene blue, Metz. 246 JOHN SUNDWALL The former stained both the epithelial cells and the granules of the mast cells a deep blue. In the latter stain the granules of the mast cells are stained metachromatically a reddish violet. In neither preparation were nerve cells and fibers observed ac- companying the blood vessels. However, claims have been made that numerous non-medullated nerves are seen in the vascular walls of the choroid plexus. Pyronin. The epithelial cells are stained deeply red. The stain is diffuse and eanfined to the cytoplasm in which the highly refractile, unstained mitochondrial granules are observed. The mast cells are not stained. No diffuse stain was seen surrounding the mast cells. This observation was made with a view of de- termining if a secretion substance is present which had been emitted from mast cells as a consequence of solution of the granules. Arnold obtained a red stained substance surrounding mast cells after vital staining with weak solution of methylene blue. This he interprets as a secretion, thus ascribing to mast cells a glandular function. Others have claimed that the gran- ules of mast cells, especially leucocytes, go into solution. I was not able to verify this observation of Arnold, either in the pyronin or methylene blue stains. NATURE AND ORIGIN OF MAST CELLS Many theories have been advanced respecting the histiogene- sis of mast cells. Lymphocytes, mononuclear leucocytes, baso- philie myleocytes and bone marrow (Schridde, ’96) are generally cited as giving origin to mast-leucocytes. That mast-leuco- eytes have a common origin along with other polymorpho- nuclear leucocytes is probable. No unanimity of opinion exists, however, regarding connective tissue mast cells (‘histiogenen mast-zellen’). That they have origin from mast-leucocytes and consequently from the same sources as the latter is he'd by many observers. Prenant, Bouin and Maillard regard the two as belonging to the same category. Ranvier (00) held that his clasmoeytes were mast cells, that they had origin from blood cells and that they were associated with inflammatory changes. Schreiber and Neuman (’01) CHOROID PLEXUS 247 agreed with Ranvier that clasmocytes and mast cells belong to the same type. Maximow states that the relation of the two—mast-leucocytes and histiogenen mastzellen—is not clear. Ehrlich held that the connective tissue mast cell is a transformed or supernourished connective tissue cell, that the granules of the latter are stored albuminous substance, and that the cells are found where tissue juices are present in abundance. Baumer (96) agreed with this conception. That the cells held a reserve substance was also claimed by Schneider (’02). Ballowitz (’96) found them to be especially numerous in hibernating animals. Others again hold that these mast cells are related to the forma- tion of fat, as they are frequently seen in adipose tissue. Unna (’91) saw the transformation of certain spindle or ellip- soidal cells into mast ce!ls in healing syphilitic ulcers. Mast cells are frequently found under pathological conditions, such as in low-grade inflammations, urticaria pigmentosa, erythema multiforme, pleuritic exudates, ete. Joachim (’06) and Spilling described mast cell leukemias. Some have made the claim that mast cells are not normally present in human blood. Michaelis (’02), Wolff (02) and many others have found them to be normally present. Audry (’96) claims that connective tissue cells, large mononuclear cells, wan- dering cells and plasma cells may become mast cells. That these cells may have their origin from blood ce'ls and vascular anlage was held by Marchand (’97). Recently much attention has been given to discussions of the origin of vascular endothelium and blood cells. That the former may develop directly from the mesenchyme is claimed by Mc- Clure (16), Reagan, Clarke (16), Danchakoff (°16) and others. Clarke (12) on the other hand maintains that lymphatic endo- thelium is formed from preéxisting lymphatic endothelium and that the mesenchyme retains throughout its identity as a mes- enchymal cell. That red blood cells develop directly from endo- thelium (even when discontinuous and independent of the cir- culation, Reagan) has been shown by the works of Reagan (16), Danchakoff ('16), Emmill (16) and Jordan (716). These observations are more or less at variance with the Angioblast 248 JOHN SUNDWALL theory of His. Stockard (15), on the other hand, did not observe in the embryo of the Teleost, Fundulus heteroclitus, any indications ‘‘that an endothelial cell has the power to produce a blood cell or to change into a blood cell of any type but much has been seen to the contrary.’ According to him, the “four distinctly different products differentiating from the apparently similar wandering mesenchymal cells’ occur under the same environmental condition. His explanation of this phenomenon is that the original mesenchymal cells that wandered out were of four potentially different classes. ‘‘The four resulting types of cells are then in an embryological sense derived from differ- ent mesenchymal anlagen,’” although these cannot be differen- tiated. The term polyphyletic theory has been applied to this conception. Against this theory is the so-cailed monophyletic theory which is ably defended by Danchakoff and Reagan. In substance it is this: There is one common anlage for all blood cells, the later differentiation into the various types of blood cells is determined by environmental conditions. To review the arguments favoring each theory would be irrele- rant in this discussion. Danchakoff, after extensive observa- tions on both the embryo and adult chicken concludes that all blood cells including mast cells, plasma cells and wandering cells have origin froma common anlage,—mesenchymal cells—and that this loose mesenchyme of a chick embryo (6-10 days) is equivalent in all the regions of localization and is polyvalent in its potencies of development. Further, Danchakoff found in the lymphatic nodes seattered in the loose connective tissue of the adult hen a loose syneytium which is considered as young un- differentiated tissue. From these loose syncytial cells amoeboid cells develop and these are the stem cells, or mother cells, of the various formed blood elements. Regarding the rdle that the stem cells play in the production of various blood cells and wan- dering cells, Danchakoff (716) states: The erythrocytes, the small lymphocytes, the different leucocytes, the wandering cells of the connective tissue, the mast cells and the plasma cells—all these cells are different cell units, morphologically as well as physiologically. But in the early embryonic stages they all had CHOROID PLEXUS 249 a common mother cell, and this mother cell is preserved in the adult organism and becomes the source of differentiation and regeneration and most probably also the source of pathological proliferation. Many diverse opinions have been held regarding the nature of the granules of mast cells. Some have regarded them as albu- minous in nature, while some regard them as mucinous. That they are associated with pathological changes in the cell and are products of inflammation or, degeneration has been maintained by others. Stoffer held that they were related to the production of melanotic pigment. Pappenheim did not regard these gran- ules as living,—‘biophoren Plasmosomen’—but a ‘substanz depot.’ He bases this view upon his observation that the gran- ules can be extruded from the cell without loosing their staining characteristics. Arnold, however, strongly maintains that mast cells are not to be regarded as cells in various stages of degenera- tion, but that they are active living normal cells. According to him, the presence of glycogen, fat, lipoids, pigment and mito- chondria is sufficient evidence of this. Others hold that the mast cell granules are secretory in nature and consequently the cell is to be regarded as a unicellular gland. The chemical structure of the granules is unknown. According to many ob- servers, they have their origin from the nucleus,—Weidenrich (11), Downey (13) and others. Arnold holds that they are related to mitochondria. Other substances found in mast cells, according to various investigators, are glycogen, fat, and pigments. Many of the theories respecting the function of mast cells have been suggested in the two preceding paragraphs. In addition to these, Fahr has held that they exert a bactericidal or antitoxic action. That the cells are to be regarded as unicellular glands is an interesting conception. This view was held by Lavdowsky, Colleya and others, who claim that the granules become disinte- grated, go into solution, and pass out of the cell to become mixed with the tissue juices, thereby contributing to the nourishment of the latter. 250 JOHN SUNDWALL DISCUSSION AND CONCLUSIONS The chief purpose of this paper is to call attention to the nu- merous interstitial granule cells in the choroid plexus of the ox, which possess the morphological and staining characteristics of mast cells of the ‘histiogenen’ or connective tissue type. Owing to the unavailableness of the ox, or the other mammals in which these cells were found in fewer numbers, for experimental pur- poses, there are many problems respecting these cells which re- main unsolved for the present. One is almost constrained to believe that their origin is from the blood, for in sections they are seen first beneath the endothelial layer of the arterioles with processes extending into the lumina; then in the walls of the blood vessels at varying distances from the Jumina, and finally beneath the epithelial layer of the choroid plexus with processes projecting between the epithelial cells into the ventricles. The entire picture suggests that the cells are trave ing from the blood vessels to the ventricles. The fact that they are seen in such variable numbers in different choroid plexuses strongly suggests this view. However, I have repeatedly attempted to find these cells in both the blood and cerebro-spinal fluid without success. It was my custom to draw off with a glass tube and rubber bulb from the ventricles, cerebro-spinal fluid whenever the oppor- tunity presented itself. About 5 ec. of fluid can be obtained in this way from the ventricles. Of course, there is some admixture of blood with this fluid, owing to the customary methods of slaughtering these animals. The fluid was then centrifugalized. I have been unable to confirm that these granular cells regu- larly pass from b!ood vessels io the ventricles, although perma- nent preparations strongly suggest such an activity. Further, I have not observed mast cells passing from blood vessels in living tissue when mounted in serum. My observations so far do not disprove the hypothesis of Ehr- lich that they are highly or over-nourished connective tissue cells. It is feasible to assume that they are wandering cells, or a special type of connective tissue cell that have become granular as a consequence of their migration through the connective tissue CHOROID PLEXUS as | wall of the choroid plexus. That they may originate entirely within the connective tissue wall is suggested in view of the fact that they were seen to undergo cell division. Certainly there is a large amount of cell nourishing substance—cerebro-spinal fluid—passing along the course of these cells from the blood vessels to the ventricles. Should we accept Ehrlich’s conception, the mast cell cannot be regarded as a specific cell, but rather as a condition of a more or less general type of connective tissue cell. Against this theory, however, 1s the fact that the formation of granules occurs simultaneously with the development of processes and that in cell division the granules are abundantly seen in the two daughter cells during the process of division. That they are not specifically related to fat formation in the choroid plexus seems certain, for fat is found in this tissue only in very minute quantities. The cells are in no way related to any demonstrab!e pathologi- cal conditions. Hence the assumption that they represent ab- normal processes is untenable. Likewise, they cannot be con- sidered as related to the formation of pigment. That the cells are concerned in some endocrinous secretion early suggested itself. Their presence in such constant and large numbers, their structure and staining characteristics, which in many respects resemble that of other gland cells, might indi- cate this. It was not until basic stains were used and the cells were seen to stain metachromatically with polychrome methylene blue that they were placed in the category of mast cells. Not- withstanding this fact, they may be concerned in some type of secretion. Many observers still regard mast cells as having a secretory function. Against this view of the secretory function of these cells may be advanced the fact that they are found only in a few mammals—in the ox, in great abundance, less numerously in the sheep, and only sparsely in swine. Further, no demonstrable physiological action was obtained after injec- tion of ox choroid extract into the dog. I was unable to demon- strate a pericellular secretion substance, as observed by Arnold. Proof is still wanting that these cells are concerned in the secre- tion of some specific substance. 252 JOHN SUNDWALL The apparent migration of these cells through the choroid plexus from the blood vessels to ventricles and the presence of inter- and sub-cellular canaliculi suggest that the cerebro-spinal fluid, at least in part, may follow such a course and consequently be independent, to some degree at least, of the surface epithelial cells. The chief value of this paper is to point out that the choroid plexus of the ox offers excellent facilities for teaching and study of mast cells. BIBLIOGRAPHY Arnotp, J.- 1914 Uber die Formbestandteile der histiogenen Mastzellen und ihrer Funktionen. Uber Plasmastruckturen. Jena, p. 344. AvpreyY 1896 Monatschr. Prakt. Derm., 22, Quoted from Enzy. der mik. Tech., II, 1910. Battowirz, E. 1891 Uber das Vorkommen der Ehrlich’schen granulierten Zellen (‘Mastzellen’) bei winterschlafenden Sdugetieren. Anat. Anz., 6, 7p: 135: Baumer 1896 Arch. Derm. Syph., 24, Quoted from Enzy. der mik. Tech., II, 1910. Benstey, R. R. 1910 On the nature of the canalicular apparatus of animal cells. Biol. Bull., 19, 3, p. 179. 1911 Studies on the pancreas of the guinea pig. Am. Jour. Anat., 12, p. 389. Craccto, ©. 1913 Uber die Anwesenheit der lipoiden Substanzen in den Mast- zellen. Zentralbl. f. Pathol. Crarke, E. R. 1912 Further observations on the living growing lymphatics; their relation to mesenchymal cells. Am. Jour. Anat., 13, 2, p. 347. See also Anat. Ree., 11, 1, p. 1. Crarke, W. C. 1916 Experimental mesothelium. Anat. Rece., 10, 4, p. 301. Cowpry, E. V. 1916 The structure of the chromophile cells of the nervous system. Contributions to Embryology, Carnegie Institution of Washington, No. 2. Cusninec, H. 1914 Studies on the cerebro-spinal fluid. J. of Med. Res., 31, Deal: Dancuakorr, VERA 1909 Untersuchungen iiber die Entwicklung von Blut und Bindegewebe bei Vélgeln. Arch. Mikr. Anat., 73. 1916 The wandering cells in the loose connective tissue of the bird and their origin. Anat. Rec., 10, p. 483. 1916 Origin of blood cells. Anat. Ree., 10, 5, p. 397. See p. 415. See also Anat Rec., 10, 3, p. 192. Dixon, W. E., anp Haturpurton, W. D. 1913 The cerebro-spinal fluid, 1, secretion of the fluid. J. of Physiol., 47, No. 3, p. 215. Downey, H. 1913 The granules of polymorphonuclear leucocytes. Anat. Anz., 44. 1913 The development of histogenesis mast cells, etc. Fol. Haemat.,16. Euruicu, P. 1877 Beitriige zur Kenntniss der Anilinfiirbungen und ihrer Ver- wendung in der mikroskopischen Technik. Arch. Mikr. Anat., 18, p. 263. CHOROID PLEXUS 253 Eneuicu, P., Kraust, R., Mosss, M., Rosin, H., Wetcert, K. 1910 Enzy- klopidie der mikr. Technik., 11, p. 69. Emmitu, V. C. 1916 The cell clusters in the dorsal aorta of mammalian em- bryos. Am. Jour. Anat., 19, 3, p. 401. Faur Ein Beitrige zum studium der Mastzellen. Arch. Path. Anat., 179, p. 450. Finpuay, J. W. 1889 The choroid plexuses of the lateral ventricles of the brain, their histology, ete. Brain, 22, Part II, p. 161. FLemmine, W. 1871 Weitere Mittheilungen zur Physiologie der [cttzelle. Arch. Mikr. Anat., 7, p. 328. 1876 Beitrige zur Anatomie und Physiologie des Bindegewebes. Arch. Mikr. Anat., 12, p. 391, 434, 500. 1879 Beitriige zur Kenntniss der Zelle und ihrer Lebenserscheinungen. Arch. Mikr. Anat., 16, p. 302. Gateorti, G. 1897 Studio morfologico e citologico della volta del diencefalo in aleuni vertebrata. Riv. di Pat. Nerv. e Ment., 12, p. 480. GOLDMANN 1913 Experimentelle Untersuchungen iiber die Funktion der Plexus choroides und der Hirnhaute. Arch. Klin. Chr., 101, p. 735. See also Miinch. Med. Wochenschr., 60, p. 1005. 1913 Vital-firbung am Centralnervensystem. Beitriige zur Phys- iologie des Plexus choroid und der Hirnhaute. Berlin, 1913. HaArckEL, E1859 Beitriige zur normalen und pathologischen Anatomie der Plexus choroides. Arch. Path. Anat., 16, p. 253. Hammar, J. A. 1895 Zur Kenntniss des Fettgewebes. Arch. Mikr. Anat., 45, p: 512. Hoyer 1890 Uber den Nachweis des Mucins in Geweben mittelst der Farbemetode. Arch. Mikr. Anat., 36, p. 310. HoumGrRen, E. 1902 Neue Beitrige zur Morphologie der Zelle. Merkel und Bonnet Ergeb. der Anatomie, 11, p. 274. Hueurnin, B. 1912 Mastzellen mit sudanophilen Granula. Zentralbl. f. allg. Pathol., 123. Joacuim, G. 1906 Uber Mastzellenleukiimien. Miinch. Med. Wochenschr., p. 1878. Jorpan, H. E. 1916 Evidence of hemogenic capacity of endothelium. Anat. Rec ONS peslye KoELLIKER, A. 1896 Handbuch der Gewebelehre des Menschen. Korysutt-Daskiewicz 1878 Uber die Entwicklung der Nerven aus Plasma- zellen beim Frosche. Arch. Mikr. Anat., 15, p. 1. LuscuKa 1855 Die Adergeflecte des menschlichen Gehirns. Berlin. Matiory 1914 Principles of Pathology, p. 24. MarcHanp 1897 Sitz. Ges. Nat., Marburg. 1898 Verh. Deutsch. Path. Ges., 1. 1901 Verh. Deutsch. Path. Ges., 4. Quoted from Enzy. der mikr. Tech., 2, 1910. Maximow, A. 1906 Uber die Zellformen des lockeren Bindegewebes. Arch. Mikr. Anat., 67, p. 688. McCriureg, C. F. W. 1916 Experimental confirmation of the view that lym- phatic endothelium arises in loco from intraembryonic mesenchymal cells, etc. Anat. Rec., 10, 3, p. 222. 254 JOHN SUNDWALL McKiesen, P. S. 1914 Mast cells in the meninges of necturus and their dif- ferentiation from nerve cells. Anat. Rec., 8, p. 475. Meek, W. 1907 A study of the choroid plexus. Jour. Comp. Neur. and Psy- chol., 18, 3, p. 286. Micuartis 1902 Uber Mastzellen. Miinch. Med. Wochenschr. Morr, F. W. 1910 Lectures on the cerebro-spinal fluid. The Lancet, July 9. MitNcHHEIMER 1895 Fort. Med., quoted from Enzy. der Mikr. Tech., 2, 1910. PappENHEIM, A. 1901 Wie Verhalten sich die Unna’schen Plasmazellen zu Lymphocyten? Arch. Path. Anat., 166. Pertir AND GirarpD 1902-1903 Sur la Fonction Secretoire et la Morphologie des plexus Choroides. Arch. d’ Anat. Mikr., 5, pp. 213-264. Prenant, A., Bourn, P., Martuarp, L. 1904 Traite d’Histologie, I, Cyto- logie, pp. 538, 595, 637. Rast 1902 Mraéek, Handb. Hautkrankh. Ranvier, L. 1900 Arch. d’ Anat. Mier., 111, quoted from Enzy. der Mikr. Tech.. 2, 1910. Ravuper-Kopscn 1906 Lehrbuch der Anatomie. 7 Auf., p. 81. Ravupnitz 1883 Centralb. Med. Wiss, quoted from Enzy. der. Mikr. Tech., 2, 1910. Reacan, F. P. 1916 Experimental studies on the origin of intraembryonic endothelium and of blood cells. Anat. Ree., 10, 3, p. 235. See also Anat. Rec., 10, 2, p. 79, 99. ScuHarer, HE. A. 1912 Quain’s Anatomy, 11, Part I, pp. 108, 128, 129, 390. ScuneEIwer, K. C. 1902 Verg. Histol., Jena. SCHREIBER AND NeuMAN 1901 Festschr. Jaffe Braunschweig. Scurippe 1896 Miinch. Med. Wochenschr. Scuutrz, M. 1856 Miiller’s Archiv, p. 311. SCHWENTER-TRACHSLER 1906 Monat. Prakt. Derm., 45. Scorr, W. J. M. 1916 Experimental mitochondrial changes in the pancreas in phosphorus poisoning. Am. Jour. Anat., 20, 2, p. 237. SPILLING 1880 Zeitschr. Klin. Med. Srockarp, C. R. 1915 2e Se ila eeaaraeeeeest ; (GSR2S5S0500005508 BODY GROWTH OF RAT 263 There were 15 test animals examined. In 10 the ovary was not diseased. In 5 the ovaries were pathological, being repre- sented by cysts. The weights of the 10 normal ovaries were as follows: one was 44 per cent less than the table value, the remaining 9 were all above the table values. On the average these 9 remaining ovaries weighed 148 per cent more than normal. They were therefore considerably over twice the normal weight to be expected from the reference tables (Donadlson, °15). It is to be noted that in the reference tables the weights are given for both ovaries taken together. CONCLUSIONS 1. Both the double and single isolation experiments show that the glandular function of the ovary which affects body growth is unmodified when the ovary is isolated from its connection with the uterus—since the isolated, acts like the normal ovary to inhibit growth. 2. This inhibition of growth is exercised in the case of ‘double isolation’ by ovaries that appear under-size or pathological at the end of the experiment (200 days), while in the case of the single isolation experiment—ovaries, for the most part greatly hypertrophied, exercise the same control. 3. From the fact that the same effect follows from ovaries in such different conditions it would appear that they probably exercise their control through the mediation of some other less modified member of the endocrine system. LITERATURE CITED Donaupson, H. H. 1915 The Rat—Data and reference tables. Memoirs of The Wistar Institute of Anatomy and Biology, No. 6. Philadelphia. Harar,S. 1913 The effect of castration, spaying or semi-spaying on the weight of the central nervous system and of the hypophysis of the albino rat; also the effect of semi-spaying on the remaining ovary. Jour. Exp. Zool., vol. 15, pp. 297-314. STrotseNBuRG, J. M. 1913 The effect of spaying and semi-spaying young al- bino rats (Mus norvegicus albinus) on the growth in body weight and body length. Anat. Rec., vol. 7, pp. 183-194. ANTERIOR HAEMATOPOESIS IN CHEMICALLY TREATED TELEOST EMBRYOS UNDER CONTINUAL OBSERVATION! FRANKLIN P. REAGAN, EDWARD E. MACMORLAND AND STUART MUDD Department of Comparative Anatomy, Princeton University ELEVEN FIGURES Inhibition of the movement of body-fluids has recently been employed for the purpose of obtaining information concerning the normal origin of teleost blood corpuscles. Those investigators who have employed this method of study have emphasized the importance of a thorough knowledge of the history of the experi- mental material from which conclusions are to be drawn. For instance, it has been stated (10, p. 579) that if an investigator knows the entire history of a given embryo, having determined during that time the embryo in question has never had any cir- culation whatsoever, he will invariably find that blood cells are located only in the intermediate cell-mass and on the posterior and ventral yolk surfaces; that there may be some variation but none “‘sufficient in any case to confuse the problem;” that erythrocytes in such embryos (8) will never be found in the an- terior mesenchyme, anterior vessels, anterior yolk surface, heart or liver, ‘‘in any embryo at any age;” that in these latter regions (9, p. 315) wandering blood anlagen never make themselves manifest. Along with these statements, no claim is made that their author has followed in sufficient detail the history of the development of any embryo to justify an assertion that the complete history of any embryo was known. Perhaps the most 1The present communication represents a portion of a work involving con- tinual observation which was aided by a grant from the National Academy of Sciences. 265 266 F. P. REAGAN, E. E. MACMORLAND AND S. MUDD definite available statement in this connection (10, p. 578) is the following: ‘“‘Since one is able to be absolutely certain that the blood never circulates in a great number of embryos, only such embryos should be considered in a study of blood origin.”’ It is evident that if one should use as his criterion for lack of circulation a corresponding lack of erythrocytes in certain loca- tions—if he should refuse seriously to consider all embryos with erythrocytes in these locations because of pictures which can be obtained in embryos by the arrest of a once established circula- tion, he is using a method capable of giving only one result. On the other hand, it has been suggested (5, p. 104) that this same thoroughness of knowledge of a given embryo’s history, such as could be obtained only by constant observation, might conceivably heighten the probability of a detection of erythro- cytes in those locations in which they have been claimed never to originate. It has been stated that circulation, after having once been established, may be lost temporarily intermittently or perma- nently. Temporary or intermittent loss of circulation would, of course, lead to error of interpretation, provided that circulation were of a sufficiently elusive nature to take place only between those intervals at which a single investigator could reasonably be expected to make his observations. It seems that the only means of meeting this difficulty of elusive circulation (if there be such a phenomenon) is to examine as often as possible the ex- perimental material. Since the working hours of a single inves- tigator might be inadequate for the elimination of the objections which might be raised by believers in elusive or intermittent cir- culation, the problem would best be attacked by the codpera- tion of a sufficient number of observers that the probability of detection of an elusive circulation approaches a maximum. It is, however, impracticable to observe with absolute constancy any single embryo. In the first place, it is impossible to predict which chemically treated embryos of a large number will sue- cumb to treatment, develop a circulation, or develop without a circulation. Since embryos of the latter sort are generally few HAEMATOPOESIS IN TELEOST EMBRYOS 267 in number compared with the other two sorts, constant watching of only one individual would necessarily involve a waste of time, and would no doubt be superfluous, even if one could be sure that a given embryo would develop without a circulation. At any rate, the method in the present work was continually to elimi- nate from a group of chemically treated embryos, those indi- viduals which developed a circulation, and those which died or became extremely abnormal. Such groups of treated embryos were carefully observed day and night at intervals varying from thirty minutes to an hour or slightly more. Low and high power binocular microscopes were employed in these observa- tions. In order that individual cases might receive adequate attention, it was necessary to limit the number of embryos very considerably below that which might be studied if observations were to be made only a few times each day. The primary purpose of the present work was to obtain a definite answer to the simple question: are erythrocytes ever found in the anterior mesenchyme or in any anterior vessels of teleost embryos in which the blood has never circulated? A negative answer to this question has been insisted upon by Stock- ard, while a positive answer has been likewise insisted upon by Reagan. The possibility of migration of erythrocytes or their anlagen is entirely beside the question. The issue concerns the presence or absence of erythrocytes in this location regardless of all possible means of attaining that location with the exception of passive movements induced by heart pulsation. Reagan (5, p. 116) has pointed out the fact that Stockard (8) has described erythrocytes as ‘originating’ on the yolk when their ultimate anlagen must have come from another location; the former has also pointed out the fact that even the leuco- eytes and their anlagen, as described by Stockard, were unable to migrate. In consideration of ultimate origins it is well to remember that the intermediate cell-mass itself arises by the gross mesial displacement and subsequent union of two longi- tudinal columns of mesenchyme. It may be stated at once that a small number of embryos which developed without circulation, as was determined by fre- 26S F. P. REAGAN, E. E. MACMORLAND AND S. MUDD quent observation, possessed erythrocytes in their anterior mesenchyme. It has previously been admitted (5, p. 112) that a large number of embryos without circulation may fail to exhibit red blood cells in their anterior regions. It was first pointed out by Reagan that blood shares with other tissues the especial sus- ceptibility of the anterior end of the embryo to a derangement of metabolism. One can readily obtain large numbers of embryos devoid both of circulation and of anterior erythrocytes; but if the middle and posterior regions of such embryos were found to be as greatly arrested and abnormal as the anterior portions gen- erally are, they would be rejected as material in which to study the origin of the vascular tissue or the origin of any other tissue. This specific behavior of the anterior tissue is included among the extensive studies by Child on Axial Gradients. To confirm the work of Stockard, one must reject all embryos exhibiting anterior erythrocytes regardless of their histories, and maintain that in the remaining embryos with diminutive anterior ends, erythrocytes have developed in all those places in which they could ever develop in the normal individual; one must claim that there is no possibility of varying degrees of haematopoetic po- tentiality. It seems unreasonable to assume that all regions should exhibit equally strong tendencies to produce blood cells, or that they should all necessarily produce them at the same early period of ontogeny. Further study alone will determine the reasonableness of this assumption. The material for the present work consisted of a few hundred embryos of Fundulus heteroclitus, which, at a very early stage, were treated with chemicals. The conditions in one of several satisfactory embryos obtained (1.e., having anterior erythro- eytes) will suffice for descriptive purposes. The known history of this embryo will be given in detail. The eggs constituting the group from which this embryo was taken were fertilized at 4 p.m., June 22, 1916. At the two cell stage, they were placed in a solution consisting of 70 ec. of sea water to which had been added 10 ec. M/12 butyric acid. Four hours later the solution was diluted by the addition of 50 ce. sea water. In this mix- ture they remained until the total time of treatment amounted HAEMATOPOESIS IN TELEOST EMBRYOS 269 to twenty-four hours, after which they were reared in sea-water which was frequently changed. From the time of their removal from the solution of butyric acid the eggs were frequently ob- served. Recorded observations were commenced on June 26 at 8 a.m., although at this time there was no sign of endothelium on the yolk-sac. Examinations were made as follows: June 26, a.m.;/9.20, 10.30, 11.45; p.m., 12.20, 1.30, 3.30, 4.45, 5.34, orkl 7.00; 8:05, 9.02, 10:00, 10:55, 11.54. . Jone 27, a.m., 1.00, 2.32, ie ro. O50; 7-50, 9:00; 10:15,-11:20° p:m., 12:10; 1.30, 2.11, 3.26, 4.19, 5.30, 6.14, 7.35,.9.00, 10.55. June 28, a.m., 12.30, 2.00. (So far there had developed no pigment on the yolks and none of the embryos possessed pericardia. Constant observation up to this time was quite superfluous.) 3.12, 4.15, 4.45, 6.00, 7.00, 8.02, 9.14, 10.25. p-m., 1.45, 3.14, 4.15, 5.11, 6.30, 8.02, 9.00, 9.55, 11.00. June 30, a.m., 12.20. At this time the embryo from which figures 1 to 11 are taken was found to contain anterior erythrocytes; this embryo was thoroughly studied, sketched and then preserved in picro-acetic acid. It might be of interest to note that none of the (approximately) one hundred individuals obtained in this same treatment had yet estab- lished a circulation. A little later, one of those embryos was found in which heart-pulsation was able to cause oscillation of the blood- corpuscles. This embryo was discarded; then twelve hours elapsed before another such embryo was found, observations having been kept as constantly as in the earlier stages. One embryo in which anterior erythrocytes (figs. 1, 9 and 11) were observed, and the history of which has been given, is of great interest. The yolk-sac was observed just before the pres- ervation of the embryo to be quite devoid of endothelium except in the most posterior region. This endothelium was readily found in section. Numerous pigment cells had developed on the yolk sac. Also there were many oil globules. On the posterior yolk were some large groups of developing blood cells which had yet shown little indication of possessing hemoglobin. They were detected by means of a high power binocular. Their color was only a little different from that of the surrounding mesenchyme, but their shape and refracting power were characteristic. In the posterior region of the intermediate cell-mass the developing erythrocytes had assumed a faint yellow color. There was not the slightest evidence that any of these cells had been, or were being, displaced by any sort of movement of body-fluids. No 270 F. P. REAGAN, E. E. MACMORLAND AND S. MUDD Erth. Fig. 1 Section through the anterior body axis and yolk sac of a chemically treated embryo from which all other figures in the present account were taken. The section shows lacunae of erythrocytes in the anterior mesenchyme. The yolk-sac is seen to be quite free from vascular tissues. Normally such a sec- tion should contain a cross-section of the heart. In this embryo the venous at- tachment of the heart is dorso posterior in the pericardial cavity. Continual observation exp. 2.53, 1916. Erth., erythrocytes; F. B., fore-brain; Y. S., yolk- sac. HAEMATOPOESIS IN TELEOST EMBRYOS 271 body-movements could be elicited from this embryo. The heart was very inconspicuous (fig. 8) and was detected with great difficulty. A very slight twitching could be observed on the ventral side of the head which gave much the appearance of slight irregular contraction of a body-muscle. The wave of contraction was directed anteriorly, appearing superficially to be quite opposite in direction from that generally found in embryos at this stage. This circumstance finds its explanation in the fact that the venous attachment of the heart to the yolk-sae is situated at the extreme dorso-posterior limit of the pericardium ventral to the axial body-tissue (figs. 8 and 10); in the normal individual the venous portion of the heart is attached to the yolk at the antero-ventral limit of the pericardial cavity. In this specimen there is no endothelial tissue near the venous end of the heart. Also there are no blood corpuscles near it, though two cells were found among the myocardial cells which had become faintly eosinophilous, and one was much rounded. The endo- cardium is represented by a very delicate cord of cells which are not sufficiently numerous to enclose a cavity (fig.6). The myo- cardium is weakly developed. Considering the fact that this embryo is less than six days old, it seems improbable that any yolk sac endothelium should have formed and then degenerated. That endothelium which could be observed in the living condi- tion was readily found in section. At any rate, continual ob- servation established the fact that there was no circulation. In the living condition the body tissue was clear and of healthy appearance. It seems quite likely that the prevention of cir- culation in this instance may have been due to an inherent ab- normality of the heart. The anterior end of this embryo was so little affected, judging at least from external appearances, that a circulation might well have developed later, provided the heart had attained iis normal relationships. It is impossible to state to what extent the chemical treatment is responsible for this heart abnormality. Embryos of this sort are very desirable for study. In many in- stances a local and apparently insignificant abnormality may be sufficient to prevent circulation in embryos whose general con- 272 F. P. REAGAN, E. E. MACMORLAND AND S. MUDD ditions are such that a circulation might well be expected. Such embryos are certainly as useful, as far as the lack of circulation is concerned, as those in which the agent of arrest of circulation has produced generalized abnormality with specific njury to the anterior tissues. It is in the former type of embryos that ante- rior erythrocytes are most often encountered. In figure 1 it will be seen that blood cells have developed in the anterior mesenchyme of the optic region, and the median dorsal fore-brain region. As has been stated, these cells pos- sessed more hemoglobin than did the more posterior ones, though the former might easily have been overlooked in the living embryo. The fore-brain tissue in this embryo appeared in the living condition to fill an abnormally large part of the head-region. The head-region itself seemed to be perfectly normal in size, differing in this character from the great majority of embryos in which circulation was prevented. In the fore-brain region, rather un- usual developmental procedure is in evidence. There are several evaginations from the brain which bear a resemblance to optic cups. On each side there is a large cup and some smaller ones. Both dorsally and ventrally the fore-brain has given off evagina- tions which in some cases are so fused with the body-wall ecto- erm (ectoderm in case of the dorsal and lateral ones) that no line of demarcation can be detected between the two ectodermal tissues (fig. 2). This hyperactivity of the fore-brain tissue is sufficiently often accompanied by the excessive development of anterior erythrocytes to warrant the suggestion that the two phenomena may be causally connected. Reagan (5, p. 107) has deseribed a case in which supernumerary prosencephalic evaginations were accompanied by anterior erythrocytes. Figs. 2 to 5 Sections through the fore-brain, showing hyperactivity of that tissue (encephalocoels). Exp. 2.53, 1916. Figs. 6 and 7 High-power drawings of sections through the heart of this same embryo. Figure 6 lies in the arterial region. The endocardium consists of a single strand of cells. Hnde., endocardium; Myc., myocardium. Fig. 7 Section through the venous end of the heart in which neither myo- cardium or endocardium is distinguishable from mesenchyme. HAEMATOPOESIS IN TELEOST EMBRYOS | 274 F. P. REAGAN, E. E. MACMORLAND AND S. MUDD In this connection the work of Child (2) is of extreme in- terest. He has found that long and severe treatment eliminates or prevents the formation of median or axial tissues, thus caus- ing bilateral structures to fuse; thet this fusion 1s most pro- nounced in the anterior-most tissues and progresses gradually backwards; that the originally distinct eye-spots of Planarians ean thus be caused to fuse into a single median one. Werber (13) suggested vertebrate cyclopia to be an expression of this same sort of procedure. As a matter of fact the vertebrate otocysts themselves may be caused to fuse into a single median one which surrounds itself with a single otic capsule. As yet it had occurred to no one to suggest that the ears arise from a median anlage, as has been maintained for the eyes. But it has also been found by Child in a work which is not yet published, that while the an- terior end suffers this specific injury with some treatments, other methods of treatment less severe and of slighter duration pre- ceding removal to the normal environment will produce condi- tions under which the anterior end recovers with greater rapidity than does the posterior end, and a condition of hyperactivity may exist. Child was able by these latter methods to produce four-eyed Planarians. These results on Planarians are paralleled recently by the unpublished work of Child on Echinoderm plu- tei. By long and severe treatment, it was found that the morphologically anterior skeletal elements may be made to fuse, while in the ease of weaker and shorter treatments, the angle of divergence between them may be made to approach 180°, while other anterior tissues become correspondingly hyperactive. While the recorded instances of anterior hyperactivity are few in the case of vertebrate embryos under treatment, it is possible that methods might be obtained whereby such hyperactivity might be obtained more often than by methods at present known tous. Quite independently of the results of Child, we have found 2s he has found in his unpublished work, that hyperactivity of the anterior end is often obtained when embryos are exposed to strong solutions for a very short time, followed by weaker solu- tions before removal to the normal environment. From such treatments, however, embryos may be obtained which appear HAEMATOPOESIS IN TELEOST EMBRYOS BS to have suffered neither arrest nor over stimulation in the an- terior tissues. While such embryos may exhibit many anterior erythrocytes, the lacunae observed there are not so large as in those embryos whose anterior ends have been overstimulated. There is one point of great importance in regard to the stain- ing properties of early erythrocytes. When these are stained with methyl blue and eosin, or when they are stained with Wright’s blood stain, the staining reaction is practically the op- posite of that of older erythrocytes. Also the staining reaction of early erythrocytes in a single embryo may be opposite or very different in different regions. In blood-smears from early embryos, such as four-day em- bryos, which seem to have been most successfully stained, the nuclei of the blood cells (all of which appear to be erythrocytes) stain reddish purple or red, while the cytoplasm appears blue or even densely blue and faintly granular; this is the picture obtained from Wright’s blood stain. In eosin-methyl blue, the nucleus stains red or purple, while the cytoplasm appears dense blue, pale blue, greenish blue or in some cases very slightly purple. These staining-reactions of course may vary with the technique, but the general results above described are most often obtained. In sixteen-day embryos, or even in stages younger than this, when the red corpuscles have well developed haemoglobin, the staining reaction is practically opposite that of erythrocytes of four-day embryos. In these advanced stages the best-stained preparations show erythrocytes with deep blue or purple nuclei and with deep red, pink or almost clear cytoplasm. If one em- ploys such stains as the haematoxylins, the nuclei of all stages can be made to stain darkly so that even with a counterstain, the method seems incapable of demonstrating the nature of this remarkable change which takes place.? *A reversal of staining reaction of nucleus and cytoplasm was observed many years ago by Auerbach. Wallin (Anat. Rec., vol. 9, no. 6) observed a reversal of staining reaction in mesenchyme cells and in free blood-cells in the anterior mesenchyme of the larva of Petromyzon. Also this result was ob- tained from ordinary stains. No doubt the entire chromatin content of a cell THE ANATOMICAL RECORD, VOL. 12, No. 2 276 F. P. REAGAN, E. E. MACMORLAND AND S. MUDD A most interesting case of variation in the staining reactions of erythrocytes within a single embryo is that exhibited by the individual which is figured in the present communication. As has already been noted, the anterior erythrocytes were more deeply red in the living condition than were those of the posterior region, the latter appearing only faintly yellow. It was pos- sible to mount enough of the sections on one slide so that the staining conditions may be assumed to be sufficiently uniform in different regions to warrant the belief that actual differences are portrayed. The nuclei of the anterior erythrocytes stained homogeneously red and purple while the cytoplasm stained pink. The erythrocytes in the posterior part of the intermediate cell- mass were found to possess red and purple nuclei of granular character and pale blue or densely granular blue cytoplasm. The intervening regions exhibited transitional conditions between these extremes. These peculiarities of the staining reactions of erythrocytes are worthy of consideration in connection with the anterior ‘leucocytes’ described by Stockard (9, pp. 280 and 281). It is interesting to note that these ‘leucocytes’ were found almost entirely in embryos about seventy-two hours old. It may be regarded as highly significant that their first recorded counter- part in the normal (?) series is found in a sixteen-day embryo, although the statement is made (9, p. 236) that the entire experi- mental series was checked by corresponding normal stages. The mere fact that these particular cells ‘‘stained differently from any other cells within the embryo” or that they are ‘peculiar’ (9, p. 280) certainly comprises insufficient grounds for their in- terpretation as leucocytes. This hiatus in the production of leucocytes between the ages of three and sixteen days is most puzzling. If it be true, as suggested by Reagan (5, p. 113) may become oxychromatic. It may, however, be doubted whether Wallin is really dealing with acidophylic chromosomes. He shows no distinct chromo- somes. It is possible that in his figures 5 and 6 he has cut diagonally through a spindle so that at one pole the section passes through a sheath of oxychromatic substance while at the other it passes through the central core of basichromatic. The latter, however, should in all cases show at least a thin periphery of oxychromatin. HAEMATOPOESIS IN TELEOST EMBRYOS Pg iT | that these are really abortive attempts at erythrocyte forma- tion, and that these cells have, or have had the potentiality of elaborating hemoglobin, it seems difficult to explain the fact that they were never detected as erythrocytes in stages slightly later than seventy-two hours unless these were then discarded on the assumption that the blood had circulated. Stockard (10, p. 578) regards all the ‘beautiful blood islands’ and ‘great clots’ in the anterior end of the embryo as mere ‘pitfalls’ to cor- rect observation. But if embryos containing anterior lacunae of erythrocytes must necessarily have had a circulation, it should still be possible to find these ‘leucocytes’ somewhere in the embryo. Diligent search for such leucocytes in embryos of this sort has many times proved fruitless. A careful study was made of some of these embryos which were able to develop circulation following the chemical treat- ment; this involved frequent observation of about twelve hun- dred embryos. In every instance in which a circulation was known to have existed and then to have stopped, the embryo failed to re-establish a circulation and soon died. Two hundred embryos, furthermore, which had established circulation subse- quent to chemical treatment were placed in a solution of 0.01 per cent potassium cyanide to which was added a rather strong solution of acetone. Examined after having been subjected to this severe treatment for one and three-quarters, five and one half, eighteen and one half, and twenty-eight hours, the embryos were found without exception to have retained their circulation. At the end of forty-eight hours only two of the two hundred embryos had lost circulation; these quickly disintegrated with- out having resumed circulation. From the foregoing results, the conclusion seems justified that the intermediate cell-mass and the posterior and ventral yolk surfaces are not the only locations in the teleost embryo which are capable of giving rise to red blood cells. Just to what extent any experimental condition represents the normal is difficult to say. The fact that a great many embryos with arrested develop- ment fail to show anterior erythrocytes might conceivably mili- tate against the belief that the anterior mesenchyme normally 278 F. P. REAGAN, E. E. MACMORLAND AND S. MUDD contains erythrocyte anlagen. It is, however, a very significant fact that the more normal the general conditions to be found in such embryos, the more likely they are to develop anterior erythrocytes. In the greater number of cases of developmental arrest, those conditions which will prevent circulation will also prevent the formation of anterior erythrocytes. It seems probable that when an embryo is once able to estab- lish a circulation, it has little tendency to lose that circulation under favorable environmental conditions. When loss of cireu- lation does occur, the embryo usually dies. We have found no case of temporary loss and re-establishment of circulation— much less, any sort of elusive or intermittent circulation. Also there has been no instance in which a pericardium was observed to become oedematous or the heart string-like, once the cireula- tion had been established. We do not assert that such things never happen. As a matter of fact, one does well to refrain from sweeping generalizations of a negative character in dis- courses dealing with the developmental possibilities of chemi- eally treated embryos. It is not our purpose to apply the facts above presented to considerations of mono- and polyphyleticism, except to state that they furnish a positive disproval of one of the contentions upon which support of the polyphyletic view has recently been based. There are certain facts which can be determined; beyond this, all is mere speculation. One can determine three things concerning the origin of a tissue: first, whether it arises from a narrowly limited anlage; second, whether any other tissue can ever give rise to the tissue in question; third, whether the as- sumed anlage never gives rise to any other tissue except the one for which it has been claimed to be specific. If the develop- ment of a given tissue fails to conform to any of these three possibilities, polyphyleticism can never be proved. There seems to be no little confusion as to the proper appli- ‘ation of the results of the study of vascular tissues in the teleost to the Angioblast theory. Stockard believes that the studies on yolk-sac endothelium by Wenckebach, Raffaele, and himself, help to disprove the Angioblast theory. HAEMATOPOESIS IN TELEOST EMBRYOS 279 Sabin (Science, August 4, 1916, p. 155) states: “In his (Stock- ard’s) studies made on the yolk-sac of the living fish embryo, he found that endothelium arises as spindle-shaped cells which differentiate out of the mesenchyme. Moreover, he found that the endothelial cell was distinct from the blood cell. This con- firmation of the angioblast of His I regard as very important.” The truth is that the observation of local formation of endo- thelium on the fish yolk-sae or on any other yolk-sac proves or disproves neither of the conflicting views. Both views admit of locally formed yolk-sac endothelium. The fact that the endo- thelial and corpuscular end-products of differentiation are dis- tinct lends no more support to the Angioblast theory than to the Local Origin theory. If they were not distinct, how could we diagnose them? ‘The statement that two differentiated prod- ucts are different is simultaneously so correct that it is redundant and so redundant that it is correct. Their distinctness does not preclude community of descent; this is a fact which we grasp with great difficulty. The real problem is whether we will recog- nize or refuse to recognize the transitional stages between the end-products, provided such can be found. Until recently, most of us have refused to recognize the transitional stages which undoubtedly exist between mesenchyme and endothelium, and now that we have done so, there is a movement to attach to individual cells in this transitional stage the appellation ‘Angio- blast,’ so that whereas there was originally one angioblast there are now as many thousands of them as the case may require. It is true that certain words in the sciences have undergone evolution in meaning; this is true of the word ‘cell,’ but this evolution did not serve to confuse the issues of a controversy. Inasmuch as Hooke did not consider a cell to be a vacuum, the term is not so inappropriate as it is sometimes considered. Stockard objected vigorously to the account of the develop- ment of the blood in Keibel and Mall’s Human Embryology for the reason that here the monophyletic view is (3, p. 350) accepted ‘with open arms’ (9, p. 310). But this account is written in strict accord with the Angioblast theory. It is difficult to recon- cile polyphyleticism and angioblast, if angioblast be a ‘unit 280 F. P. REAGAN, E. E. MACMORLAND AND S. MUDD anlage’ (3, p. 499) of which all vascular tissues are ‘direct descendants’. There have appeared recent discussions over blood-histogenesis in which authors would seem to believe themselves to have settled by words the entire question of preformation—a feat which Bonnet, about two centuries ego proclaimed as ‘‘une des plus belles victoires que ’entendement put ait remporte sur les sens.” LITERATURE CITED (1) Bonnet, C. 1762 Cont. dela nat., 7 me partie, c. ix; Oeuvres, vol. 4, p. 270. (2) Cutty, C. M. 1912 Certain dynamic factors in the regulatory morpho- genesis in Planaria dorotocephala in relation to the axial gradient. Jour. Exp. Zodél., vol. 13. (3) Kerpet, F., anp Mati, F. P. 1912 Manual of Human Embryology, vol. 2. J. Lippincott Co. (4) RarragLe, F. 1892 Ricerche sullo sviluppo del sistema vascolare nei Selacei. Mitth. a.d. Zoolog. St. z. Neapel, Bd. 10. (5) Reacan, F. P. 1915 A further study of the origin of blood vascular tis- sues in chemically treated teleost embryos with especial reference to haematopoesis in the anterior mesenchyme and in the heart. Anat. Rece., vol. 10, no. 2. (6) 1916 Experimental studies on the origin of intraembryonic endothe- lium and of blood cells. Proce. Am. Assn. Anat., vol. 10, no. 3. (7) Reacan, F. P., ann THorineton, J. M. 1915 The vascularization of the embryonic body of hybrid teleosts without circulation. Anat. Rec., vol. 10, no. 2. (8) Srockarp, C. R. 1915 An experimental study of the origin of blood and vascular endothelium in the teleost embryo. Proc. Am. Assn. Anat., Anat. Rec., vol. 9. (9) 1915 The origin of blood and vascular endothelium in embryos with- out a circulation of the blood and in the normal embryo. Am. Jour. Anat., vol. 18, no. 2. (10) 1915 A study of wandering mesenchymal cells on the living yolk sac and their developmental products: chromatophores, vascular en- dothelium and blood cells. Am. Jour. Anat., vol. 18, no. 3. (11) Wenckrepacnu, K. F. 1885 The development of the blood corpuscles in the embryo of Perea fluviata. Jour. Anat. and Physiol., vol. 19. (12) ISS5S Beitriige zur Entw. der Knochenfische. Arch. fiir. Anat., Bd. 20. (13) Werper, BE. 1. 1915 Experimental studies aiming at the control of defec- tive and monstrous development. Anat. Rec., vol. 9, no. 7. PLATE 1 EXPLANATION OF FIGURES | 8 Sketch of the living embryo of which all other figures in this account rep- resent sections or portions thereof. The erythrocytes in the head, tail and on the posterior yolk are represented by dots. The entire yolk sac except the posterior surface is devoid of blood and endothelium. There are large oil- globules on the yolk. Exp. 2.53, 1916. 9 Portion of a section through the fore-brain region in which the hyper- active brain-wall has fused with the ectoderm, enclosing on all sides some mes- enchyme which became haematopoetic. There is no endothelium present. If any has been present it has either disappeared or turned into erythrocytes. F. B., fore-brain; Ect., ectoderm; Erth., erythrocytes. 10 Section through the body-axis of the same embryo showing the loose mesenchyme (mch) in which the venous end of the heart loses itself. 282 HAEMATOPOESIS IN TELEOST EMBRYOS F. P. REAGAN, E, E. MACMORLAND, AND S. MUDD PLATE 1 PLATE 2 EXPLANATION OF FIGURES 11 Portion of a section through the fore-brain region of the embryo from which the previous figures were made. It was drawn at high magnification. Lying free in the mesenchyme are erythrocytes apparently in various stages of development. The epithelium with large cells and dark intercellular substance is prosencephalic tissue. 284 HAEMATOPOESIS IN TELEOST EMBRYOS PLATE 2 F, P, REAGAN, E. E. MACMORLAND AND S. MUDD OESTRUS AND OVULATION IN SWINE GEORGE W. CORNER AND A. E. AMSBAUGH From the Anatomical Laboratory of the University of California I. THE PERIOD OF OVULATION It seems remarkable that almost nothing should be known of the process of ovulation or of the mature ovum of the pig, which has for many years been in constant use for the investigation and teaching of mammalian embryology. In so thorough a review as Marshall’s ‘Physiology of Reproduction’ (10) there is only a meagre note: It is probable that the sow ovulates during oestrus and not during the pro-oestrum, since it is stated that sows are most successfully served on the second or third day of ‘heat.’ Coition, if it occurs earlier, is frequently not followed by conception. From Hausmann’s description (1840) it would seem that ovulation does not take place prior to coition, but this statement has not been confirmed. That ovulation occurs at or near the time of oestrus is stated in Keibel’s Normentafeln (97) and is of course implied in Assheton’s studies on the early development of the pig (’98), but the precise relation of the two events has never been fully worked out. To obtain exact information upon this point we have under- taken the observations here reported. They have been made possible through the generous cooperation of Mr. J. O. Snyder, of the Western Meat Company, San Francisco,! and of Mr. Ralston B. Brown, Superintendent of the Oakland Meat and Packing Company’s plant in West Berkeley, and his staff. Through Mr. Brown’s kindness permanent laboratory space had 1 We hope to have an opportunity to express fuller thanks to Mr. Snyder and the gentlemen of the U. 8S. Inspection service at the South San Francisco plant, in connection with other studies to appea™ somewhat later, the material for which has been obtained through their immediate assistance. 287 288 GEORGE W. CORNER AND A. E. AMSBAUGH already been provided at the packing house; for this work we were allowed free access to the stockyards at all hours, and upon occasion animals weve killed out of the regular order for us. The females of the wild swine of Europe are monoestrous, according to Kaeppeli (08), having but one period of heat in the year; but under domestication the sow becomes polyoestrous, coming in heat at intervals of two to four weeks, usually about every twenty-one days, as all breeders agree. The period of heat. commonly lasts three days and is characterized by sexual excitement and in some individuals by swelling, reddening, and slight eversion of the vulva, or even at times by a serous, mu- cous, or partially sanguineous discharge from the genital ori- fice. If a boar be present, the sexual excitement is made ap- parent by ready acceptance of coitus, which is denied at all other times; if none but females are in the pen, the sow in heat will be seen to sniff at the genitals of her neighbors and ‘ride’ them in imitation of coitus. Frequently the sow is the recip- ient, rather than the donor, of these attentions. The period is not terminated by coitus, but continues until the end of three days. In order to distinguish these sows from others in the corrals until the time of butchering, they were marked with a daub of white paint thoroughly rubbed into the hair of the back between the shoulders. Immediately after evisceration, the Fallopian tubes were removed by cutting across the upper portion of the uterine horns, were carried to the laboratory in 0.7 per cent saline solution, and there washed out by inflating them with salt solution through a slit in the wall near the fimbriated ex- tremity. After inflation with the fluid, the tubes were gently ‘milked’ into a Syracuse dish, and the washings examined with the dissecting microscope. This simple and almost infallible method of finding the ova was suggested to us by Professor Evans as an improvement on Martin Barry’s practice of milking the tube without injected fluid (’39). As we have subsequently found it had been used by Sobotta and no doubt others as well. Our series includes ten animals which were in heat on the day of killing or the day before. In eight of these the Graafian OESTRUS AND OVULATION IN SWINE 289 follicles had ruptured and we were able to recover some or all of the ova from the tubes in each case. Of these eight sows, six were killed on the second or third day of the period, one between sixteen and thirty-nine hours after the onset of heat, and one between thirteen and twenty-two hours after the be- ginning of oestrus. In two of the ten there were large Graafian follicles, all unruptured except one follicle in one of the sows, which had apparently just collapsed. Unfortunately we have no record as to the time of onset of heat in these two animals, but the conditions in the other eight show that ovulation had occurred during oestrus, and probably on the first or second day of the period. II. OVULATION SPONTANEOUS IN THE SOW During the discussion which arose over Born’s and Fraenkel’s suggestion that the corpus luteum exercises the function of in- ducing ovulation at regular periods, a distinction was drawn between those mammals in which ovulation is spontaneous, and those in which copulation is necessary to invoke rupture of the follicles. Villemin (’08) maintained that ovulation is spon- taneous in all mammals, but Ancel and Bouin (’09) state, on the basis of personal researches (details of which are not given) that ovulation is spontaneous in the human species and in other primates, in the dog, horse, cow, and pig; in the rabbit, guinea- pig, mouse, and cat rupture ensues only after coitus. The work of Marshall and Jolly (06) on the dog and Heape (’97) on the mare, are in agreement with the results of Ancel and Bouin, and to the first mentioned class we may also add the sheep (Mar- shall ’03) and the rat, according to Sobotta and Burekhard ('10) confirmed and extended by the recent carefully gathered data of Long and Quisno (’16). The placing of the rabbit in the second class has been confirmed by Regaud and Dubreuil ('08) ; the cat by Longley (°11); and Marshall (04) has added the ferret to the list. The mouse, however, belongs to the class in which ovulation is spontaneous (Tafani ’89, Sobotta ’95) and also the guinea-pig (Loeb 711). 290 GEORGE W. CORNER AND A. E. AMSBAUGH The only mention of the sow in this regard is the statement of Hausmann (’40) quoted above from Marshall (10) that in this species ovulation is not spontaneous. On the contrary, our specimens show clearly that in swine coitus is not necessary for rupture of the Graafian follicles, for we have records of ten sows in which ova were found in the tubes although no boars had been in the pens with them. Mr. Brown informs us that ac- cording to the conditions of shipment of the live-stock, it is very unlikely that these animals had the opportunity to copulate before arriving at the stockyards. Two of the sows were under observation before oestrus set in, and are therefore even more definitely known not to have copulated. Moreover, in an animal in which but one follicle of many had ruptured, copulation had been observed sixteen hours previously. Ovulation, therefore, is independent of copulation. Ill. THE MATURE OVUM Little or nothing has been known of the mature ovum of the sow, and we have found no record of any previous observation of the unsegmented ova from the tube. Assheton’s earliest specimens were already in the two-cell stage (98). Lowrey (11), in his study of the prenatal growth of the pig, attempted to estimate the size and weight of the mature ovum by allowing a slight addition to the diameter of the largest ovarian ovum he found, which measured 177 micra, with a zona pellucida 10 micra in thickness. He estimated, therefore, that the mature ovum would have a diameter of 180 micra. We have measured four- teen fresh tubal ova from nine sows, and find the diameter, in- cluding the zona pellucida, to vary from 155 to 165 miecra, the zona being about 10 micra in thickness. The ova are plainly visible with the naked eye if placed against a strong light. We have not noticed a radial striation of the zona pellucida either in fresh or fixed ova. The ovum is filled with yolk granules of varying sizes, usually about three to five micra in diameter, which are so numerous and so refractile that they quite conceal the nucleus. A polar body may often be seen very clearly. OESTRUS AND OVULATION IN SWINE 291 The ova are usually naked, but may be covered by the cells of the corona radiata, or even by a considerable portion of the discus proligerus. A few of the ova which have been sectioned, seem to show no deviation from the usual process of maturation in other mam- mals; the first polar body and the second polar spindle are formed in the ovary, and the second polar body seems to be formed after fertilization. In each of two sows killed on what we believe to be respectively the sixth and eleventh days after the onset of oestrus, we were surprised to find a degenerating ovum in the tube. In three sows in which copulation had occurred, fertilized ova were found. ‘They all chanced to be in the same stage, just be- fore fusion of the two pronuclei, and are therefore the earliest embryos of the pig yet reported. The ovaries and uteri of these animals are naturally of the greatest interest, and studies of them are now in progress. SUMMARY 1. In the domestic sow, ovulation occurs during oestrus, prob- ably on the first or second day. 2. Ovulation is independent of coitus. 3. The mature unfertilized ovum of the sow measures 155-165 micra in diameter, has a zona pellucida about 10 micra thick, and a yolk heavily laden with fat. 4. Fertilization of the ovum occurs in the Fallopian tube, as in other mammals. THE ANATOMICAL RECORD, VOL. 12, No, 2 292 GEORGE W. CORNER AND A. E. AMSBAUGH LITERATURE CITED ANcEL, P. AND Bourn, P. 1909 Sur les homologies et la signification des glandes & séerétion interne de lovaire. C. R. Soc. de Biologie. Paris, T. 67, p. 464. AssSHETON, R. 1898 The development of the pig during the first ten days. Quarterly Journal of Microscopical Sciences, vol. 41, p. 329. Barry, M. 1839 Researches in embryology. Philosophical Transactions, pt. 1, 1839. Hausmann, U. F. 1840 Ueber die Zeugung und Entstehung des wahren weib- lichen Eies. Hannover (quoted by Marshall, 1910). Heapr, W. 1897 The artificial insemination of mammals. Proc. Royal So- ciety, vol. 61, p. 52. Kareprett, I. 1908 Beitrige zur Anatomie und Physiologie der Ovarien von ' wildlebenden und gezihmten Wiederkiuern und Schweinen. Berne, dissertation. Keiser, F. 1897 Normentafeln zur Entwickelungsgeschichte des Schweines. Jena. Lors, L. 1911 The cyclic changes in the ovary of the guinea-pig. Jour. Morph., 22, p. 37. Lone, J. A. AND Quisno, J. E. 1916 The ovulation period in rats. Science, N.S., vol. 44, p. 795. Loneitey, W. H. 1911 Maturation of the egg and ovulation in the domestic cat. Am. Jour. Anat., vol. 12, p. 139. Lowrey, L. G. 1911 The prenatal growth of the pig. Am. Jour. Anat., vol. 12, p. 107. MarsuHatu, F. H. A. 1903 The oestrous cycle and the formation of the corpus luteum in the sheep. Philosophical Transactions of the Royal Society, B, vol. 196, p. 47. MarsuHauu, F. H. A. 1904 The oestrous cycle in the common ferret. Quar- terly Journal of Microscopical Science, vol. 48, p. 323. 1910 The Physiology of Reproduction. London. MarsHatu, F. H. A. ano Joutyy, W. A. 1906 Contributions to the physiology of mammalian reproduction. Part I: The oestrous cycle in the dog. Philosophical Transactions of the Royal Society, B, vol. 198, p. 99. ReGaup, Cri. anp Dusreutn, G. 1908 L’ovulation de la lapine n’est pas spon- tanée. C. R. Soc. de Biologie, Paris. T. 64, p. 552. Sopnorra, J. 1895 Die Befruchtung und Furchung des Eies der Maus. Archiv fiir Mikroscopische Anatomie, vel. 45, 8S. 15. Soporra, J. AND BurckHarp, J. 1911 Reifung und Befruchtung des Eies der weissen Ratte. Anatomische Hefte, 42, 8. 433. Tarant, A. 1889 I primi momenti dello svilluppo dei mammiferi. Atti d. R. instituto di stud. super. prot. e. di perfezion, Firenze. (Quoted by Sobotta 95). VitLeMIN, F. 1908 Sur les rapports du corps jaune avec la menstruation et le rut. C. R. Soe. de Biologie, T. 64, p. 444. INTRA-UTERINE ABSORPTION OF OVA ARTHUR WILLIAM MEYER From the Division of Anatomy of the Stanford Medical School SEVEN FIGURES While collecting embryological material for other purposes a decade since, my attention was not infrequently arrested by the presence of a degenerating or retarded sheep embryo in an apparently normal uterus the other horn of which contained an apparently normal foetus. Although these embryos were not infrequently quite normal in form they were always decidedly smaller than the normal embryos. In most cases the conditions suggested pathological changes both within the uterus and the embryo. The amniotic fluid was sometimes intensely turbid and even milky and more frequently dark in appearance and very evidently contained degenerating blood. Since twin pregnan- cies are not so very frequent in sheep, the number of cases seen was necessarily small. In uteri containing normal foetuses in the early stages of development, the embryo, the development of which had been arrested, was often represented by a rather firm fleshy mass of regular form which sometimes showed unmistak- able evidences of degeneration even upon inspection with the unaided eye. While engaged in the determination of the curve of prenatal growth in the guinea pig, Draper and myself found several cases of abnormal or at least regressive ova. All these abnormal guinea pig ova were much smaller than the normally developed ones of corresponding age should have been. Indeed, most of them were represented by firm oval fleshy masses, some of which possessed protuberances which made them look bicornuate. All were fastened with one end in more or less distinct uterine crypts which were especially evident in one of the smaller ova. The 293 294 ARTHUR WILLIAM MEYER instances met with so far were seen in pigs killed 19, 25 and 37 days after coitus. In the first case that of guinea pig No. 16 only a single ovum was found present twenty-five days after coitus. This ovum was contained in the distal extremity of the right horn and was surrounded by a reddish black fluid. The uterus and adnexa appeared wholly normal to the naked eye. ‘The ovum was com- posed of a slightly oval fleshy mass only 5 mm. in diameter al- though the normal embryo of this age measures 17 mm. It had a smooth regular surface and was still attached to the opened uterus but was easily detached. No placenta or foetal mem- branes were recognizable and the ovum protruded freely into the opened uterine cavity being attached to the uterine mucosa by its base with its longest diameter perpendicular to the latter. The line of attachment on the ovum apparently formed about one-eighth of its total perimeter. On sectioning, the tissues of this ovum were found to be de- cidedly degenerated, the outer layers being composed of nothing but cell detritus. A little beneath the surface, this cell detritus is mixed with degenerating mesenchyme and variously-sized, better-preserved epithelioid cells. Between the latter he large numbers of erythrocytes. These are scattered about freely and occupy other areas almost exclusively. Polykaryocytes and megakaryocytes in various stages of degeneration and different forms of leucocytes are also present. Some of the giant cells contain very bright golden pigment some of which is found also extra-cellularly. The framework of degenerating embryonic connective tissue, contains scattered cells and groups of cells with extremely large vesicular nuclei and prominent nucleoli. Deeper beneath the surface remnants of blood vessels and of a reticulum which reminds one of that in young lymph nodes can be seen. In some areas, however, nothing but the degenerating reticulum with a little granular detritus remains. In addition to the giant cells large irregular masses which look like fused giant or other cells are also seattered through the specimen. Sections made through the middle of the ovum show that the portion nearest the area of contact with the uterine mucosa is INTRA-UTERINE ABSORPTION OF OVA 295 best preserved and composed of a syneytium-like mass in which large vesicular nuclei predominate. This portion also contains a large vesicle lined by a low embryonic epithelioid syncytium. The spherical vesicle which in its largest portion comprises more than one half the diameter of the ovum contains nothing but a transparent fluid. Similar much smaller vesicles are also scat- tered about throughout the rest of this portion of the ovum some lying isolated at its very perimeter. A similar low epithelioid layer with indistinguishable cell boundaries also covers a portion of the surface of the most degenerated distal portion of the ovum where it also clothes villus-like extensions from the main mass. Some of the sections are almost surrounded by this epithelioid layer. Small areas of these sections are practically devoid of tissue and contain almost nothing but a faint reticular network enclos- ing a slightly granular detritus and many polymorphonuclear leucocytes the nuclei of which have a typical horseshoe shape and the protoplasm of which is acidophile. Some of these leuco- cytes look decidedly degenerate and none seem to be phagocytic. In other often adjacent areas, the place of the polymorpho- nuclear leucocytes is taken by somewhat large cells with a vesicu- lar nucleus which is circular in outline. The protoplasm of many of these cells is acidophile but here and there groups which look bright golden are seen. Most of these cells are well-preserved but some of them can be seen to be filled with similarly staining erythrocytes and what look like fragments and granules of erythrocytes. Specimen No. 17 taken from a pig pregnant 19 days contained three ova, a normal one in the left horn and two abnormal ova in the right. The normal embryo weighed 35 mgm. and the smaller of the abnormal ova was approximately as large as the placenta and membranes of the normal embryo which weighed 0.91 mgm. The larger ovum was bicornuate. Both were single masses and no distinct placental portion was recognizable. Both these ova which were no larger than a normal embryo of this age, were regular in form and their surfaces smooth. Upon microscopical examination, however, a few small, villus-like ex- 296 ARTHUR WILLIAM MEYER tensions were seen on the distal portion. A few small indenta- tions were also evident but nothing else interrupted the regu- larity of the rest of the surfaces. The larger of these two ova was very well-preserved and much more vascular than that from pig No. 16. It was covered throughout by a low epithelioid syncytium which evidently was originally composed of a low cubical epithelium for here and there cell outlines are still faintly visible or the free surface of the syncytium is indented quite regularly so as to look crenated (fig. 1). These crenations are evidently the result of projections formed by the individual cells with the indentations located in the region of the former cell boundaries. The slightly irregularly-shaped, evenly-staiming nuclei are thickly packed and although the cell boundaries are not clearly recognizable the layer is low and in places contains indistinct lines which look like remnants of cell boundaries and justify one in characterizing the cells as cuboidal. The rest of the ovum is composed of a syncytium containing large vesicular nuclei as shown in figure 2. Cell boundaries are distinet no- where but the tissue apparently was a large-celled mesenchyme originally. Only a few small areas of almost complete degenera- tion are present. The tissue is densest and least vascular near the region of attachment to the uterine wall. The most rarefied tissue is found in the two cornua and near the distal portion where the loose mesenchyme contains small bloodvessels. Im- mediately beneath the investing epithelioid layer the specimen is completely canalized by wide capillaries which form an ex- ceedingly vascular peripheral layer. A bit of the less vascular portion is shown in figure 3. This specimen contains no large cavities but numerous smaller epithelioid-lined spaces are found in the cornua and the distal Fig. 1 Structure of a portion of the periphery of a nineteen-day ovum. X 475. Fig. 2. Structure of a portion of the periphery of another ovum of the same age. X 515 Fig. 3 Structure of the vascular portion of the ovum shown in figure 2. x 475 Fig. 4 Large nuclei from the central necrotic area of one of these ova. * 475 INTRA-UTERINE ABSORPTION OF OVA 298 ARTHUR WILLIAM MEYER portions. The largest cavity which is lined by a rather low de- generated epithelioid layer is found in the center of the speci- men but it is so small that it is not visible with the naked eye in the stained section. Although better preserved these cavi- ties or cysts are similar to the very large one contained in the previous specimen. In spite of the condition existing in this specimen the blood is all contained in vessels only a few of which can be distinguished as veins or arteries. Some of the small arteries which are located in the basilar portion of the ovum near the uterine attachment have become completely obliterated. The portion of the ovum near the area of attachment is almost non-vascular as in the previous specimen. In some of the outer portions, however, and also near the placental attachment the tissue is of a more fibrous nature and looks far less embryonic. The blood cells are well-preserved and all the leucocytes have round vesicular nuclei in contrast to those found in specimen 16. The second specimen from No. 17 was somewhat smaller and without cornua but it also was surrounded by an abundantly nucleated syncytium and was canalized beneath its surface by numerous capillaries as shown in figure 1. Portions of the sur- face were also pitted by crypts which gave the periphery of the sections a fenestrated appearance. All these crypts and vesicles are lined by a similar syncytial layer and all are empty. The specimen like the previous one is most vascular near the surface and near the fenestrated portion where the tissue composing it is also much looser. Only a few villus-like processes are seen in the distal portion. Although half—apparently the proximal half—of this speci- men was lost, the structure of the remaining half is practically the same as that of the previous ovum. Its preservation is not quite so good, however, for it contains partially necrotic and small liquefied areas in its interior. The more necrotic por- tions of this ovum contained nuclei truly gigantic in size. This will be evident on comparing the magnification of figure 4 with those in 1 and 3. It too is quite vascular and some of the ves- sels all of which are full of blood, are extremely large. Some portions of this ovum look more like fibrous mesenchyme others INTRA-UTERINE ABSORPTION OF OVA 299 more like sarcomatous tissue, but cell outlines are nowhere evident. Specimen 19 in which the period of gestation was twenty-six days contained five abnormal ova, two in the left and three in the right horn. All of these were quite equally-sized, irregular masses but they were only about two-thirds as large as a normal placenta with that duration of pregnancy. They were easily detached and projected freely from the opened uterine cavity as had those in the previous cases. In all except one ovum the placental crypts were very shallow fossae but this specimen was contained in a definite funnel-formed uterine erypt about 3 by 3mm. insize. Since these crypts were not noticed until the ova had been removed from the uterus I am inclined to think, how- ever, that they were formed mainly by the post mortem contrac- tions of the uterine musculature. This assumption is also sug- gested by the fact that all these ova completely fitted the lumen of the uterus and formed slight elevations on its surface. No pla- cental portion was recognizable with the naked eye and there was no gross evidence of pathological changes in the uterus. Although these five specimens of abnormal ova varied some- what in size this variation was not marked. All were from 4 to 6 mm. long and 2 to 4mm. thick and in contrast to the pre- ceding specimens the four ova which were removed from the uterus had a dull fuzzy instead of a smooth shiny surface. They were exceedingly soft and rather irregular in shape. The contracted uterus which looked entirely normal was nodular in consequence of the enlargement opposite the ova. It contained no exudate and upon microscopical examination the ova were found well-preserved. All these specimens were but. slightly vascular, the small capillaries being located mainly in the peripheral layers as before. They were all devoid of an outer epithelioid layer and were composed of a syneytium containing large nuclei none of which were nearly as large as those found in the preceding specimens, however. As in the previous specimen the largest nuclei were found in the interior of the ova and the smallest at the surface where they were more elongated and where the syneytium took on a more 300 ARTHUR WILLIAM MEYER fibrous and stratified character because the tissues and the long axes of the nuclei, were arranged parallel to the surface. In one portion of one ovum the extremely large cells with their large oval nuclei are still preserved and give one a good idea of what the original structure of the ovum, in the early stages of: de- generation really was. No other type of cell was found except that the formation of the giant cell masses is indicated through coalescence of ad- jacent degenerating cells. The capillaries are engorged with erythrocytes and a few leucocytes with vesicular nuclei, but no vessels larger than capillaries of the ordinary calibre are present anywhere. The structure of these ova at the region of the uterine attachment corresponds to the rest. One of these specimens still shows a little of an epithelioid covering in two very small places. In one of these the epithe- lium is shown in the form of a tube which may represent the remnant of a erypt or an invagination. Although this ovum is completely canalized by a plexus of fine capillaries near the periphery it contains no larger vessels in its interior. One of these five abnormal ova from No. 19 was left in situ and cut serially in paraffine. It measured 5.5 mm. in diameter after fixation and completely filled the uterine cavity except in a few areas where small spaces were left between the ovum and the uterine wall. t 7 if . ia . ey An: | us t a 1 x rn ‘ a ‘ i 4 ia | eae dt! Se Bere! * Pate tn ) ca a v ow ve ny hn’ n 4 4% ae eK® ‘ : : ny ¢ ss x. : i : ye eae | aE eet ae. eh ene hi Fa 1 + ret } ‘Stel Mata sia eree sted ys; | Gah Sony on Pe ; - ms i ; ny ae , ed era oe Py Mant ei “Sey TSE - Bag ‘by Waste eee ee hi y Fe, 4 Pf METHODS OF MOUNTING SECTIONS IN GELATIN J. B. JOHNSTON AND EDNA G. DYAR University of Minnesota Gelatin has been used to some extent by various European workers for mounting sections, but has not come into general use. Our experi- ments show that gelatin possesses three properties which render it undesirable for this purpose until overcome by special treatment. Gelatin is non-hydroscopic, brittle and inelastic, and instable. In the extremely dry winter climate of Minnesota the gelatin dries and shrinks until it bends or breaks the glass slide on which it is spread, or cracks and peels off the slide. In very moist summer weather the gelatin may become whitish-opaque from moisture. When these qualities are corrected by the addition of glycerin to render the film hydroscopic, sugar to make it elastic and pliable and some hardening agent to render it stable, a satisfactory and permanent mounting medium is secured. To render the gelatin insoluble in water, formalin, chrome alum, chromic acid, tannic acid or potassium bichromate may be used with varying results. The most satisfactory product is obtained by means of potassium bichromate or chrome alum. After the addition of one of these salts the gelatin is rendered insoluble upon exposure to light. There is also produced a very slight grayish-green tint by the chrome alum and a deeper greenish tint by the bichromate, but this does no harm in thin films. Perhaps the best results are obtained by pro- tecting the gelatin by means of an insulating varnish. The varnish seals the gelatin against the action of atmospheric vapor and also prevents the evaporation of water from the hardened gelatin film, the presence of water being necessary to maintain elasticity and pliability. The advantages of a gelatin mounting medium are: saving of expense of dehydration for mounting in balsam and of the cost of cover glasses, and availability in some cases where an aqueous medium is necessary. It can be used on sections stained with haematoxylin, carmine and some but not all of the anilin dyes (e.g., not with acid stains soluble in water). Its use in films without glass may prove to have some value. SOLUTIONS TO BE EMPLOYED A. Best quality photogelatin 5 grams. Distilled water 100 ce. Add glycerin 5 ec. Let stand two hours. Raise to 50°C. Gelatin dissolves. Filter through canton flannel. 309 310 J. B. JOHNSTON AND EDNA G. DYAR B. Hydrate 2 grams, gelatin in 45 ce. distilled water. Add 10 ce. glycerin and 15 ce. corn syrup. Raise to 50°C. When gelatin is dissolved add 18 grams gelatin hydrated and dissolved in 55 ee. distilled water. Filter through canton flannel. C. Prepare as in B, using 2 grams gelatin and 45 ec. water and 23 grams gelatin and 55 ce. water. D. To render A, B, or C insoluble add to the solution prepared as above 14 ee. of a 10 per cent solution of potassium bi- chromate or chrome alum to every 100 cc. of the gelatin solution. This solution must be used at once, since it will not melt after _ being allowed to harden. In making the above solutions a temperature of 45° to 48°C. is necessary to dissolve the gelatin. After the gelatin is dissolved the solution may be lowered to 30°C. without causing the gelatin to set. In mounting sections it is necessary to keep the gelatin sufficiently warm to secure penetration, but advantage should be taken of the fact that the gelatin remains fluid at lower temperatures which are less likely to harm the tissues. In the following directions it is intended that the gelatin itself shall be kept at about the temperatures indi- cated, whether by means of an oven or a constant temperature plate. For paraffin sections Bring into distilled water on the slide. Place slide on constant: temperature plate at 35°C. and cover with sufficient solution A to make a complete covering film when dry. After a few minutes on the warm plate set in a horizontal position to harden. For celloidin sections up to a thickness of 100 microns Carry the sections on paper (‘onion skin’ best). Clear in glycerin and water over night, followed by glycerin several hours. Immerse sections in solution A in flat dish at 35°C., 20 minutes. Clean and flame! a slide and place on warm plate. Spread on slide a small amount of solution A and immediately place section on it, remov- ing the paper. Carefully press out all air bubbles. Add more solution A and then drain to secure a thin but complete covering for the section. Keep on level warm plate until gelatin is evenly spread. Place in horizontal position to dry at room temperature. ‘Flaming the slides is necessary to remove the thin film of organic material which is taken up from the air by slides exposed for any length of time. The usual cleaning solutions do not wholly remove this. MOUNTING SECTIONS IN GELATIN aut To secure a sufficiently thick and even covering, when the five per cent gelatin is used it is best to make the first coat thin and apply a second and third coat to the slide when cold, draining each time. For thicker celloidin sections (1 to 2 mm.) After clearing in glycerin immerse the sections for at least three hours in solution A at as low a temperature as practicable (30°C.) and mount in solution B by the method just given. For mounting sections in films without glass Fasten together two very thin celluloid films by means of snaps on one edge, place the section or sections between them and immerse in water or glycerin at 35°C. Drain, separate the films and sections as they are immersed in solution B in a flat dish on warm plate. Abun- dance of gelatin solution must be used, to secure complete immersion. After twenty minutes carefully press out the air bubbles with the fingers or a rubber wedge. Attach snap hangers to all four corners of the film and hang up to dry at room temperature. The hangers at the lower end serve as weights to prevent warping. For thick sections (1 to 2 mm.) in films, immerse first in solution A at least three hours and then mount as just directed, using solution C. As this thick solution does not drain off readily, the excess gelatin should be removed from the surface by stripping the film between thumb and finger. Solutions A, B, and C may have thymol added and be kept cold and remelted when wanted. Finally, in each of the above cases either an insoluble preparation (solution D) should be used or the finished preparation should be ecov- ered with a varnish, as soon as it is dry enough not to be sticky. One may use the Zapon varnish found in the market. The same may be made by dissolving 5 grams celluloid in 20 cc. amyl acetate and 80 ce. acetone. A good varnish which is more pleasant to use is made by . Abney as follows: Alcohol 20 ounces Ether 40 ounces Pyroxylin or celloidin 400 grains In applying the varnish care should be taken to have the gelatin com- pletely covered. Films should be dipped in the varnish, and on slides the varnish should be spread with brush or spray beyond the edges of the gelatin. December 8, 1916 THE VALUE OF ABSOLUTE ALCOHOL FOR REMOYV- ING ADHERENT PARAFFIN SECTIONS FROM PAPER OR PASTEBOARD TRAYS LOUIS H. KORNDER! From the Anatomical Laboratory of the Northwestern University Medical School Laboratory workers in embryology, histology, and neurology, and especially those whose work calls for the mounting of serial paraffin sections, have all at some time been annoyed by the adhesion of their sections to the paper or pasteboard tray on which they had been placed prior to mounting. This occurs, of course, most frequently during the warmer months of the year or where sections are not mounted for a long time; thus leading often to a complete loss of a valuable series. Because of this it may be of value to call attention to the use of absolute alcohol as a means for overcoming this difficulty. The amount of alcohol used is very slight, being just sufficient to moisten the sections completely and in addition overrun on the sides so as partially to impregnate the paper or pasteboard to which they adhere. After this, several minutes time should be allowed for evaporation of the aleohol. This requires only a brief period, but can be hastened if a current of cool air is allowed to strike the tray. Where the sections adhere with especial firmness this last is particularly desirable. It is of advantage when dealing with large sections to take a section- lifter or fine-bladed scalpel and run it along under the edge of the sections before the alcohol thas completely evaporated. After this procedure sections which adhered firmly loosen almost always with ease. The use of 95 per cent alcohol is not recommended since with it satisfactory results are less often obtained. It does not evaporate with the rapidity and thoroughness of the absolute alcohol. The latter vaporizes very readily and to this its action of loosening paraffin sections from paper or pasteboard trays may possibly be ascribed. 1 Contribution No. 47, November 1, 1916. 312 PRESERVATION OF ANATOMIC DISSECTIONS WITH PERMANENT COLOR OF MUSCLES, VESSELS AND ORGANS A SUPPLEMENTARY NOTE, DESCRIBING ANOTHER METHOD, THE CURING METHOD EDMOND SOUCHON Professor Emeritus of Anatomy, Tulane University of Louisiana Since the printing of the original manuscript in the Anatomical Record, Vol. 10, No. 1, November, 1915, I have found out a new and simpler method, the curing method. It consists in the following: After the completion of the dissection, if the muscles present a dark brown color, the preparation is immersed in G33C1 for three days. G33C1 means a solution composed of 33 per cent of glycerine, 1 per cent of carbolic acid and 66 per cent of water. It registers 10° Baumé. At the end of the three days it is taken out and exposed to the air in a room until the muscles become black. This requires about ten days. If some muscles do not blacken as fast as the others, they should be painted daily with pure glycerine until they become as black as the others. As soon as all the muscles are black the preparation is immersed in Ch.F75, 1.e., a solution of chloride with 75 per cent of formal, as previously described. Then it is placed permanently in Ch.F5. The results are quite satisfactory. The advantage of this method is that it does away with the use of calcium chloride, described pre- viously, which is not always uniform. Pale muscles do not do so well by this method. They should be painted in preference. I have also brought out the following points which are of some assistance in the work. In using tallow for distending the arteries, it is best to use beef tallow from around the kidneys. It is obtained from the butchers at the market. Tallow from corn fed beef is the best. When the dissection is completed, if the muscles present a dark brown color they are suitable for the curing method. They are comparatively scarce. If the muscles show a lighter color than dark brown they are suitable for the paint method. They are much more common than the dark brown. 313 314 EDMOND SOUCHON In the final preservation of curing preparations Ch.F5 will do as well as Ch.F.20; thus effecting a marked saving in the cost of the permanent solution. I found that the following mixture of paint gives better results: Tuscan red, half teaspoonful; turpentine, 2 teaspoonfuls; lamp black, 2 grains. The lamp black tones down the Tuscan red when too bright in the solution. This quantitiy is more than enough to paint two or three times all the muscles of the upper extremity. Try the mixed paint on 1 or 2 inches of a muscle to see how the color will show and modify the paint accordingly. When using lamp black, any dark red paint will do. Tuscan red is not so essential. Two or three thin coats of paint are better than one thick coat. Thick coats make preparations look like daubs. When the preparation is placed in the solution, if the color is not satisfactory, too light or too bright, do not hesitate to take it out, expose to the air for a few hours to dry and then repaint, with a suit- ably prepared paint. Do this two or three times if necessary. Final success depends on it. Painted preparations do better decidedly on A10F.5 than in A10 alone. They do not require filtering as often and filter better. The F.5 prevents the bacterial cloudiness. It does not seem to affect the color of the paint any more than A10 or A20. Artists’ oil paints (Winsor and Newton) are better for the vessels than any other. Deep Vermilion for the arteries and Permanent Blue for the veins. When the paint remains in the cup over night, even with a lid, it becomes too thick and turpentine must be added to it before using it. A flat brush of fine bristles about 4 inch wide is quite handy to paint large surfaces. Keeping solutions clear all the time is the foundation of the preser- vation of the preparations. When solutions have been filtered twice in succession, if they do not come out clear, they should be changed for fresh solutions. Solutions very cloudy or very discolored filter badly. They clog the filters. It is best to make new fresh solutions. When solutions get old (two or three years) they should be changed unless they remain clear. THE DEVELOPMENT OF THE HUMAN CHIN W. D. WALLIS Fresno, California Linneus is generally given the credit of having been the first to make the observation that the chin is a characteristically human trait In none of the anatomical or anthropological dis- cussions of the chin has the writer seen credit for this shrewd observation given to any earlier scientist. Observation and record of the fact that the chin is a characteristically human trait is, as a matter of fact, centuries older than Linneus, and it is doubtful whether the latter did not copy it from some older authority. Pliny in Book XI, Chapter 60, of his Natural His- tory, informs us that ‘‘no animal, with the exception of man, has either chin or cheek bones.’’ The Greeks when in the act of supplication, touched the chin to show, as some would say, their affinity with the divine, and, if this is true, making fitting recognition of its human peculiarity as a trait not shared by the animals. But no Greek scientist seems to have speculated about its origin. The attempt to explain the evolution of this anatomical fea- ture is certainly comparatively recent. It appears to be no earlier than Cuvier. Cuvier remarks that in certain quadrupeds, individuals occasionally are born with the upper jaw unusually small; as a result the lower jaw, by the inturning of the alveolar processes to articulate with the unusually short upper ones, gives rise to a mental prominence bearing a close resemblance to the human chin. He claims to have seen an instance of this in a calf of Geneva, of reputed human paternity. Cuvier’s explana- tion was a shrewd one. Sir Ray Lankester has used a precisely similar argument in accounting for the evolution of the chin of the elephant which, as he has shown, has developed pari passu 315 316 W. D. WALLIS as the jaw has shortened, the unsupported upper lip mean- while lengthening into the trunk of the modern elephant. In the past century considerable attention has been given to the development of the chin and various explanations of its evolution have been offered. Any discussion of the problem should give some review of these views and we have attempted to summarize them below. SPEECH Attention has often been called to the effect, or supposed effect of speech upon the mandibular conformation—or vice versa Osborn, for example, reminds us that the narrow passage between the alveolar processes which, in the simia, lie in almost parallel, or, in some cases, in forwardly converging lines, give small range for the action of the tongue.! Whether this restric- tion of the play of the tongue would seriously interfere with speech, may be doubted. We speak of tongues wagging, but they do not really wig-wag so much as hump themselves while oscillating upward and downward in interfering with expiration. It is only upon failure of speech that we stick our tongues in our cheeks. The uplift in the human palate might be consid- ered more favorable to speech than the increased width of the alveolar processes. Defects in the palate cause defects in speech, but we do not hear that people with long, narrow jaws have less linguistic facility than those with short, broad jaws. The effect of the mandible upon speech has not, however, elicited so much defense as its converse, the effect of speech upon the mandible. In the mandible of simia will be founda deep pit lying in the inner concave surface. This seems to be absent generally in other mammals, including mankind. It accommodates the genio-glossus muscle which rises here and spreads out, fan-like, along the middle line of the lower surface of the tongue. This muscle, Dr. Louis Robinson believes? aids ‘ Men of the Old Stone Age,.100, 139-140, Scribners, 1915. ? Evolution of the chin. North American Review, Sept., 1914, vol. 200, p. 438-449. DEVELOPMENT OF THE HUMAN CHIN ° oe the tongue in sorting out the contents of the mouth. The dog, for example, seems to have considerable difficulty in getting rid of undesired morsels, while cattle, the giraffe, and the camel, shift the undesired portions to one side, where long, projecting papillae help to work them out along the cheeks as they are agitated by the tongue. In place of this pit we find in the human jaw a bony prominence or tubercle, and Dr. Robinson professes to be able to trace the gradual evolution of this pit for the accommodation of the genio-glossus muscle, as we find it in the simia, to the bony prominence known as the genial tuber- cle which replaces it in the human jaw. The pit itself is subject to much variation, being most shallow in a fossil lemur, and shallow in the anthropoid apes, because a downward tilting of the margin of the jaw below the incisor teeth gives larger surface for the attachment of the muscle. The depression seems to be least in the siamang gibbon, while the human jaws of Heidelberg and Naulette are said to show it to a slightly less extent than the siamang gibbon. As these are among the oldest and most simian of human jaws the correspondence is not to be lightly passed over. Klaatsch® confirms the existence of a fossa sub- lingualis in the Heidelberg jaw, and says that in the Hauser skull from Le Moustier no genio-glossus spinal prominence is present, its place being taken, as also in the Mauer and Krapina G. skulls, by a fossa or pit for the insertion of this muscle.‘ The insistence of Robinson is not without precedent. In 1904 Dr. C. Toldt, of Vienna, at a meeting of the German Anthropo- logical Society of that city, suggested that the evolution of the chin was due to the development of the muscle emanating from the tongue and necessitated by speech. He returned to this argument the following year, in a paper entitled “Uber die Kinn- knéchelchen und ihre Bedeutung fiir die Kinnbildung beim Menschen.’’> This paper called forth a reply by von Hause- 3 Zeitschrift fiir Ethnologie, 41 (’09), 554-555. 4See also Hausemann, ib., p. 719, 721; R. R. Schmidt, Die Diluviale Vorzeit Deutschlands, 234 (Stuttgart, 1912); K. Gorjanovic-Kramberger, Der Diluviale Mensch, 176-181. 5 Vienna Anthropologische Gesellschaft Metteilungen, 36 (’06), 51, 54. 318 W. D. WALLIS mann, who insists that the ossicula mentalia are to be otherwise accounted for, and that other causes have been primarily respon- sible for the evolution of the chin.® ; C. C. Blake? considered the absence of the genial tubercles in the La Naulette jaw purely adaptive: ‘The relative and absolute great thickness of the jaw at its symphysis originates this shelf- like structure, which is solely caused by the great deposit of osseous matter around the site of the genial tubercles.” Mr. Roosevelt finds the jaw of the chinless homo heidelberg- ensis so primitive that it must have made his speech thick and imperfect.’ The veteran French anthropologist, Paul Topinard, is not impressed by such arguments. In his discussion of the significance of the absence of the genial tubercles in the La Naulette jaw’ he points out that only a small portion of the muscles from the tongue, namely, the geniohyoid muscle, which reaches from the small hyoid bone at the base of the tongue to the symphysis, is attached to the lower part of the tubercles, the genio-glossus muscle finding attachment to the region above this. Instead of accepting the presence of tubercles as an advantage, he insists that the depression in the gorilla’s man- dible gives him considerable advantage over man. Albrecht found the mandible of an idiot of twenty-one years of age possess- ing tubercles that attained the extraordinary eminence of 9 mm.!° This outvies the tubercles of educated orators. This idiot, observes Topinard, did not have half the persuasive elo- quence doubtless enjoyed by the La Naulette lady. ® See his paper on Die Bedeutung der Ossicula Mentalia fur die Kinnbildung. Zeitsch f. Ethnol., 41 (’09), 714-721; see also P. Bartels in Int. Monatschre f. Anat. and Phys., vol. 21, p. 179 ff.; Osborn, op. cit., 228; R. Virchow, in Zeitsch f. Ethnologie, 14 (’82), 287; Schaaffhausen, in Korr. d. Deutsch Gesse f. Anthrop. (Jan. 1881), No. 1, p. 3; L’Anthropologie, 4 (’93), 753-754. 7 Anthropological Review (’67), vol. 5, 295-302. ® National Geographic Magazine, Feb., 1916. * Rey. d’ Anthropologie (’86), serie 3, vol. 1, 416-25. 19 See Bull. Soe. Anthrop. d. Bruxelles, i (’82-’83). DEVELOPMENT OF THE HUMAN CHIN i 319 DECREASE IN THE SIZE OF TEETH AND OF THE ALVEOLAR PROCESSES When our forebears assumed the erect posture and were able to fight freely with those fists which no longer had to serve as supports when running, the dangerous canines became shorter, and there was a gradual degeneration in the size, if not in the number also, of the teeth. The large bony alveolar processes in which they lay embedded were no longer needed, and there was a corresponding shrinkage in this bony structure. The absorp- tion of the alveolar region leaves the lower portion of the man- dible, which is more solid and less liable to change, relatively prominent and suggestive of a chin. This view has been popular. Toldt, in the paper referred to, insists that the reduction in the size of the teeth, together with the drawing in of the enfolding alveolar processes, would tend to develop the chin. His commentator, von Hausemann® recog- nizes the force of this portion of Toldt’s argument. Bardeleben" pointed out that the building up of the chin would result from reduction in the size of the teeth, and declared he knew of no exception to this correlation. In accounting for the reduction, he not inaptly likened the building of the chin to a hillock left out-standing on a plain where erosion has reduced the general level. The osseous portion which is left outstanding, like this older stone formation, becomes a protruding chin. The receding of the lower jaw is not to be forgotten. In this recession the teeth and the alveolar processes are especially involved, so that the protuberantia mentalis gradually comes to the fore—as we find even with prehistoricman. Similarly, Arthur Keith, while attributing to the muscles of the tongue a tendency to give a forward development to the chin, lays more stress on the reces- sion of the alveolar processes that accompanies reduction in the size of the teeth.” Robert Munro in 1912" attributed the prominence of the chin to retrocession of the facial bones, ‘‘as the shortening of 1 Anatomischer Anz., 26 (’05), 107. 12 Man: A History of the Human Body, 193-194. No date. 13 Paleolithic Man, 19 (Macmillan 712). 320 W. D. WALLIS the alveolar ridges would cause the teeth to assume a more upright setting in their sockets.”” Walkhoff noted the tendency of the reduction in teeth and alveolar processes to give rise to a chin.'* Weidenreich viewed the progressive development of the chin as purely a passive process: it got ahead by remaining where it was, the superior alveolar region being meanwhile in retreat. Rudolf Martin is inclined to champion these views'® as is also Osborn. The latter adds to this tendency the growth and specialization of the muscles of the jaw and tongue employed in speech, though he insists that absence of the chin does not betoken inability to speak. Prof. T. T. Waterman has recently added his support to this school.!? That absorption of the alveolar processes will leave the chin prominent is proved in the changes that take place in old age, where both process and result can be observed.!8 Thus the changes that take place during the life of the indi- vidual are to some extent an epitome of those that are recorded by prehistoric evidence. ‘‘In all modern races of men the front part, of the semicircle arch of teeth has shrunk or ‘withdrawn’ considerably, or more than has the bony jaw in which the teeth are set. Consequently the bone projects in front of the teeth as the bony chin.’’!® Robinson recognizes the argument that shrinkage of the alveolar processes gives rise to the chin, but discountenances it. ‘4 Die menschliche Sprache in ihrer Bedeutung fiir die funktionelle Gestalt des Unterkiefers. Anat. Anzieg., 24 (’03), 129-139; Beitrag zur Lehre der mensch- lichen Kinnbildung, ib., 25 (’04) 147-160; see L’ Anthropologie, vol. 15, 99-100, 235-236 (’04). Similarly Frizzi, Archiv. f. Anthrop, (’10), vol. 37, p. 255 ff. ‘° Die Bildung des Kinnes und seine ungebliche Beziehung zur Sprache. Anat. Anzeig., 24 (’03-’04), 545-555. ‘© Lehrbuch d. Anthropologie, 873-874, Jena, 1914. 17 The evolution of the chin. American Naturalist, April, 1916. ‘8 See the description and illustration E. G. Norris, Human Anatomy, 65. Much valuable material will be found in the article by Ernst Frizzi, Unter- suchungen am menschlichen Unterkiefer mit spezieller Beriicksichtigung der Regio mentalis. Archiv. fur Anthropologie, 37 (’10), 252-286. Contains exten- sive bibliography, 101 sketches, and detailed measurements of 100 mandibles of different races. 19 E. Ray Lankester, Diversions of a Naturalist, 250-252 (London 1915). DEVELOPMENT OF THE HUMAN CHIN 321 OTHER OSSEOUS CHANGES The changes in the anterior portion of the mandible, if we eompare that of an anthropoid with that of man, are not ex- hausted in the differences already noted, but include other important features. The os mentale in the simia is narrow, regular in curvature, and of smooth surface. The human os mentale often has a jagged border, and may be medially con- cave both in a horizontal and in a vertical plane, when viewed anteriorly. Most striking of all, it possesses, on both the right and the left side, a small bony protuberance or ossicle, which gives roughness to the contour, adds to the impression of greater breadth, and breaks up the convex lines into horizontal concave surfaces. Where normal osseous changes are not located at articular surfaces, we usually have to account for them as due, either to some shift in adjacent bony tissue which directly affects them, or to the action of muscles. The importance of the tuberosities at the symphysis in giving prominence to the chin, should not be forgotten. Their importance as a factor in the development of the chin has been recognized by B. Adachi.2?° Mies points out the value of the ossicula mentalia in building up the chin, and also the fact that paired muscles are associated with paired ossicles, whereas an undifferentiated muscle is asso- ciated with the undifferentiated ossicle.t A similar argument was later adduced by Toldt, who attempts to show that the development of the ossicula mentalia, which proceeds along different lines in the human and in the simia, has been influ- ential in giving rise to the chin.” MUSCULAR AND MECHANICAL FORCES That these bony protuberances are directly related to muscle development is more than probable. Bardeleben’s extensive investigations show that man alone possess on the os mentale 20 Uber die Knéchelchen in der Symphyse des Unterkiefers, Zeitsch. f. Morph. und Anthrop., (’04) 7, 369-372. 21 Uber die Knéchelchen in der Symphyse des Unterkiefers vom neugeborenen Menschen. Anat. Anzeig., ('93), 8 p. 361-356. 22 Die Ossicula mentalia und ihre Bedeutung fiir die Bildung des mensch- lichen Kinnes. Sitz. d. Kaiser. v. Akad. d. Wiss. (’05), 114, (AB. 111), 657-92. 322 W. D. WALLIS the paired (paariger) muscle, the musculus anomalus menti, which is associated, whether as cause or effect, with this bifur- cation of the os mentale in the human. Not only is there this difference between simia and homo sapiens, but, as a consulta- tion of G. Rugge’s, Die Gesichtsmuskulatur der Primaten” will show, considerable differences in the muscular system of the two are represented in this portion of the face. If the hand is held on the collar bone, the chin tilted in air, and the skin covering the chin raised as far as possible, the hand will detect movements over the surface of the clavicle. These movements are effected by a large muscle, or system of muscles, which rises in the lower lip, passes over the os mentale, down the front of the neck, and over the clavicle and some of the upper ribs. In the prosimia a large bundle of platysma pass over the os mentale, due partly to the narrowness of the anterior margin, partly to the demand for larger muscles in the larger lips which do much more heavy work than the human lips. The tendency wherever muscles pass over a bony surface and pull against it, is to flatten that surface. Hence the regular rounded contour of the os mentale in the simia, with absence of outstanding bony prominences such as we find in the human mandible. There is, moreover, in the human, a specialization of muscular development which has proceeded far beyond that of the apes. The os mentale is traversed also by a set of muscles known as the musculus mentalis, which run at almost right angles to the platysma. In the human face these systems are separate and specialized, while in the simia they are often so interwoven as to make it impossible to entirely separate them. In Ateles, for example, it is difficult to distinguish parts of the mentalis from the platysma, while in the chimpanzee the complication is even more marked. Similarly, according to Robinson, there is both greater development and greater specialization in the genio- glossus muscle in human beings then is to be found in the apes. “In man the genio-glossus has become a series of a large number of independent muscular strips which are to all intents and pur- poses separate muscles, each with its little fiber of the hypo- 23 Leipzig, 1887. DEVELOPMENT OF THE HUMAN CHIN 323 glossal nerve entering it in such a way as not to hamper its _ free movement, while in the apes it is apparently a single muscle, or a closely united group, acting en bloc.’’ It has been already mentioned that the paired muscle on the os mentale is found only in human beings. and that it seems to be related to the external genial tuberosities peculiar to the human os mentale. Rugge noticed that the mental region of the human skull is distinguished from that of the simia by the possession of numerous tubercles which must be supposed to be the points of origin for many small muscles not to be found in the apes. We need only consider the comparative range and facility of facial expression in the two species to grasp both the fact and the explanation of the existence of these numerous tubercles that cover the human mandible and their absence in the simia. The rapid pull of facial muscles used in articulation may have con- tributed not a little to this result. Some of the muscles used in such facial twitchings as laughter, for example, involves, find attachment on the anterior surface of the mandible. Here the bony prominences that rise to give these muscles attachment help in the outer construction of the chin. No one who has read the detailed account of the muscles used in facial expression, given by Prof. Arthur Thomson in his Anatomy for Art Students, can be skeptical about the much greater specialization of facial muscles in the human being and the tendency of these to elicit bony prominences on the skull and the mandible as points for attachment.” It is not improbable that this difference in muscular pull will do much to explain the simian type of chin. The thick-lipped peoples, as notably the Negroes, have, of course, larger muscles to work their larger lips, and they have less prominent chins. Those negroid peoples who have thinner lips have more promi- nent chins. The chin is well developed in the Eskimo, though they possess a long and heavy mandible, and large teeth. In the typical negro the chin is but feebly developed, in keeping with *4 An. Rep. Smith. Inst., 1914, p. 603; Knowledge (London), Nov., 1913. *5 See also Alfred Fripp, Human Anatomy for Art Students. THE ANATOMICAL RECORD, VOL. 12 No. 2 324 W. D. WALLIS the heavy, thick, protruding lips, though the negro mandible is not larger, longer, nor heavier, nor are the teeth larger than those of the Eskimo. In the more orthognathous Bushman, with smaller, thinner lips, we find, on the contrary, a well devel- oped chin of the anteriorly concave type. There is, in fact, a general, though not a complete, correlation between thick, promi- nent lips and retreating chins. The protruding teeth which are associated with the retreating chin, enable the animal to get rid of the food with comparative ease. In fact, if the teeth were set upright, as are ours, to dis- gorge would be fraught with considerable difficulty. If this advantage of being able to get rid of unwelcome and retain de- sired contents is to be preserved, large, long, strong upper lips are needed in a land where many hungry mouths linger round to pick up a fallen morsel or snatch a disappearing one. Our remote ancestor, ‘probably arboreal,’ had to keep a tight under lip until he could carry his head as high as the descendant who became lord of all he surveyed. Then large under lips were no longer necessary. In our contemporaries they serve merely to proclaim the proximity of their simian ancestry. Mr. G. F. Scott Elliot asserts that in consequence of the broadening of the skull and the shortening of the face into a shorter, rounded-arch shape, great strain and cross-tensions would be thrown on the extreme forward points of the jaw- bones.2° In this I do not follow him, for it seems to me the tendency would, if anything, be the opposite. Nor do I follow him in his suggestion that if the origin of bone-forming tissue may be due to muscular stresses and strains, then the produc- tion of bone-forming tissue at the chin would be favored—unless he means such stresses and strains as we have indicated. So far as mastication is concerned, the muscular stress would be much greater in the simia, which use their heavy, protruding - lips in lieu of free hands, to pull in their food, and certainly when eating, use them much more than we use ours. But the masticatory muscles are not attached to the chin and could searcely affect the os mentale directly. Simia must do more *6 Prehistoric Man and His Story, 76-77; (London, 1915). DEVELOPMENT OF THE HUMAN CHIN aap severe work with canines and incisors than we do with ours, but the muscular strain comes much further back on the mandi- ble than Elliot supposes.?7 The muscles used in deglutition must, of necessity, be larger and stronger in the simia than in human beings, and, so far as they attach to the anterior part of the mandible, we might expect them to exert some influence on its conformation. ‘That these muscles do exert an influence on the shape of the chin is more than probable. Bijvoet, in a detailed study of the morphology of the musculus digestricus mandibulae in the zoological world, including man and the primates, demonstrates its varying area and method of attachment, which is not the same in those ani- mals which chew with a scissor-like motion in the vertical plane, as in those whose jaws move with a side to side grinding motion—differences which are to some extent typified in man and the monkey, Duckworth’s assurance to the contrary notwithstanding.”® CHANGES IN HEAD FORM The lower jaw is not anatomically a part of the skull, yet it would be wrong to suppose that we can consider it as a feature isolated from the remainder of the head, since, physiologically, it is an integral part of the head. The upper alveolar processes are the supplementary portions which make, with the mandible, a functional unit. The various portions of the skull are so closely interrelated, either structurally or functionally, that any considerable change in a given region is apt to be reflected by °7 The method of muscle attachment has been well shown by C. Toldt, Der Winkelfortsatz des Unterkiefers beim Menschen und bei den Siugetieren und die Beziehungen der Kaumuskeln zu demselben. Sitzungsberichte der Kaiserlichen Akademie der Wissenschaften, 1905, vol. 114, (Abteilung IIL), 315-478. Further descriptive and illustrative account of the differences between the human and the simian will be found in C. Toldt’s papers, Der vordere Bauch des M. digas- tricus mandibulae und seine Varietiiten beim Menschen; and, Der digastricus und die Muskeln des Mundhoéhlenbodens beim Orang. ; Sitzungsberichte der Kais. Acad. d. Wiss., 1907. Ab. 111, vol. 116, p. 373-459; vol. 117 (’08) p. 229-324.) *8 Bijvoet, Zeitsch f. Morph. und Anthrop., 61 (’07-08), 249-315. W. H. L. Duckworth, Anthropology and Morphology, vol. I (15). 326 W. D. WALLIS corresponding or compensating changes throughout the entire skull.2° The upper alveolar region is an incorporated portion of the facial skeleton, and is dependent for its evolution upon changes in the facial region. The face is, in turn, architecturally but a pendant portion of, the upper supporting skull case, so that changes in the skull case, affecting the points of juncture with the facial bones, facilitate changes in the facial proportions and with these the conformation of the alveolar regions. Dr. Karl Gorjanovic-Kramberger insists that an intensive con- struction of the chin is possible only through a change in the angle of prognathism, and reduction in length and size of teeth. The change in the angle of prognathism throws the roots of the front teeth to the fore and bony tissue must be built up there to give them strength. With this change in the angle of the teeth the mental prominence becomes more pronounced and the con- struction of the chin is assured.*® He alleges also, and with some show of reason, that the construction of the chin in modern man has been directly correlated with the arching of the prognathous mandible. Toldt, similarly, has attributed the formation of the chin to progressive changes accompanying the bine = of the face and the arching of the mandible.*! The changes that have taken place with the assumption of the erect posture and the use of softer food, have been fairly uniform, and are seen in many portions of the skull. As the 29 This has been ably demonstrated by Mr. Francis Knowles in his study of the correlation between the interorbital width and other measures and indices of the human skull. Journ. of the Roy. Anth. Inst., 41 (’11), 318-49. Herman Welcker, in an article on Die Zugehérigkeit eines Unterkiefers zu einem be- stimmten Schidel, ete. (Archiv. f. Anthropologie 27 (’01-’02), 37-106, has ably demonstrated the interdependence of the mandible and the skull. See also Aurel Von Torok, Uber Variationen und Correlationen der Verhiltnisse am Unterkiefer; Zeitsch f. Ethnol, 30 (’98), 125-182. The correlation is, of course, not absolute. Exceptions exist, but the reference to them as ‘disharmonic’ types is evidence of the rule. 0 See the section on Zur Bildung des Kinnes beim Homo primigenius, in the author’s Der Diluviale Mensch von Krapina in Kroatien, 171-176, (Wiesbaden, 1900). 31 Corresp. Blatt. d. Deutsch Gesells fur Anthrop., 35, (’04), 94; see L’Anthro- pologie, vol. 16, p. 583-584. DEVELOPMENT OF THE HUMAN CHIN 327 foramen magnum moves forward and the head becomes better balanced, less muscular pull is required to keep it in position. The mastoid muscles degenerate, pressure along the temporal bones decreases, the forehead emerges, and the head increases in breadth. The face, at the same time, broadens, because its foundation walls in the calvarium have begun to shift laterally. The alveolar processes must shift laterally also. This lateral spread of the palatal region arches the frontal portion of the alveolar region, which tends to be pointed or straight in the simia, and draws the incisors in, so that they no longer project, as hitherto. The face projects less, and with the straighter face the teeth, which in both simia and homo sapiens follow the facial angle, approximate the vertical; for the teeth must conform to the plane of the bony tissue in which they are set. Since the mandible is useful only as a correlative portion of the facial structure, it must conform to the changes effected in the skull. In doing so the chin becomes, of necessity, more and more prominent. In the transition from the simian to the human type, we must take into account also the change that has come about in the articulation of the incisors. In the simia the incisors meet. This is not uncommon in palaeolithic man and in some of the more prognathous people. Among ourselves it still occurs, but only in a small percentage of cases. In the gorilla, indeed, the upper canines actually overlap the lower to a marked degree, so that the diastema on the lower alveolar region is much more marked than that on the upper. Already, then, the relative prominence of upper and lower incisors seems secured by this forward grasp of the upper canines. Man is standing proof of the triumph of the facial canines over the mandibular, for in homo sapiens all of the upper incisors, as well as the canines, easily overlap the lower, though sometimes the reverse occurs. It must, moreover, be borne in mind that the anatomical inde- pendence of the mandible, even where there is no physiological independence, allows the lower alveolar processes to accommo- date themselves more rapidly to the new conditions, and we might expect that they would be more rapidly influenced by changes 328 W. D. WALLIS in diet than would those of the palatal region. This would account for their more rapid retreat, while at the same time the enclosing upper incisors help by means of pressure from without. We do, as a matter of fact, find that: the changes in the man- dible have gone further than those in the facial portions. In primitive man the alveolar processes are still at the outer margin of the lower facial region. But not so in the jaw of civilized man with prominent chin, where the alveolar processes lie behind the underlying heavy bony tissue, so that the teeth no longer conform to the plane of the bony tissue in which they are em- bedded. We do not find such considerable changes in the facial portion as in the mandibular. We conclude, then, that no one factor should be singled out and given the credit for having evolved the chin, but that many forces have contributed to this result. Widening of the dental arcade gives more play for the tongue, whether for shifting food or consonants. It makes a better masticator, and perhaps, a better linguist. The interrelation may be close. Which is cause and which effect may be difficult to determine, for we have, not one, but many, parallel or interrelated developments converging in that peculiar human feature, the chin, which muscular forces, both within and without. have helped to design. NOTES ON TWO CASES OF ANOMALOUS RIGHT SUBCLAVIAN ARTERY RICHARD W. HARVEY University. of California Medical School Cases of anomalous right subclavian have been reported oceasionally, but their rarity and interest from an embryological point of view, besides their possible clinical relations, warrant their addition to the anatomical literature. Holzapfel (’99) collected two hundred cases of abnormal right subclavian artery including four of his own, and dis- cussed them from anatomical, developmental, and practical stand- points. He gave the frequency as one case to one hundred sixty-seven cadavers, or .6 per cent. Cobey (’14) gives the results of ani nvesti- gation for the Anatomical Society of Great Britain and Ireland as five to five hundred, or 1 per cent. The cases here reported are two to two hundred thirty-seven cadavers, or 0.8 per cent. Bean (’04) re- ported two cases of his own and six others in the literature besides Holzapfel’s cases. Cobey and Bevier (’15) have reported each one case. Case 1. The subject is a white male, age sixty years. The arcus aortae gives off three branches, the truncus bicaroticus, arteria sub- clavia sinistra, and arteria subclavia dextra, the anomalous branch. It extends in a gentle curve upwards, backwards, and to the right from the level of the lower border of the first left costal cartilage to the left side of the body of the third thoracic vertebra. This abnormal course of the arcus aortae is produced by the rotation of the base of the heart towards the left. The aorta ascendens, therefore, lies ven- trad and to the left of the aorta descendens, both lying to the left of the vertebral column. The truncus bicaroticus is flattened ventro- dorsally, and, of course, lies to the left of the trachea and the midline. The A. subclavia sinistra is the second branch. The third branch is the anomalous A. subclavia dextra arising from the right side of the aorta descendens at its commencement. It crosses the vertebral col- umn behind the oesophagus and trachea, diagonally cephalad and to the right, lying on the body of the third thoracic vertebra. It is con- siderably dilated up to the point where it emerges from behind the oesophagus. The great veins are normal. The trachea is normal. The oesophagus shows an interesting abnormality by making a detour to the right for a distance of 10 mm. where the anomalous A. sub- clavia dextra emerges dorsad to it. The right vagus preserves its usual course in the thorax. The N. recurrens is absent, the cardiac branch 329 330 RICHARD W. HARVEY of the vagus, usually arising from the recurrens, being supplied directly from the vagus trunk. Three or four twigs, taking the place of the recurrens, pass from the nerve trunk directly to the larynx. The left vagus is normal in its course and branches. The N. recurrens, however, passes across the ventral aspect of the anomalous subclavian. The ductus thoracicus is normal in its course. The phrenic nerves are normal. Case 2. The subject is a white male of advanced years: The arrangement of the arcus aortae and its branches is similar to that described in Case 1. However, the truncus bicaroticus is cylindrical, and the abnormal A. subclavia dextra is not dilated. In Holzapfel’s cases 33 were dilated out of 51 in which that feature of the abnormal artery was discussed, or 64 per cent. One of the cases collected by Bean was dilated. The clinical effects of a dilated abnormal artery ase to be taken into account, therefore. Holzapfel discusses this point and concludes that only by an aneurysmal enlarge- ment of the artery may dysphagia lusoria be considered. Surgically the abnormal course of the artery is important in attempts at ligation; in one instance reported it offered a decided difficulty. In surgical conditions involving the oesophagus or in operations on the thorax its presence may be of considerable consequence. The internist should be interested in the possibility of its causing unequal radial pulsations. Holzapfel believes it is not the anatomical cause of left- handedness, although it is to be noted that the two conditions exist in several cases. Cobey suggests the possibility of the abnormal artery producing symptoms similar to those of cervical rib, with resulting trophic changes in the extremity. LITERATURE HouzapFeL, G. 1899 Anat. Hefte. Bd. 12, p. 369. Brean, R. B. 1904 Johns Hopkins Hosp. Bull., 15, p. 203. Copry, J.F. 1914 Anat. Rec., 8, No. 1, p. 15. BevierR, G. 1915 Anat. Rec., 9, No. 10, p. 777. GH SA BPAT A IB Bye HT He | fo RUE KE dy na AE RA REE CRORE ARR ING ORT MRED BE ABR Seah S HA carr? soy ma= HR K HA SH @ WK Bh WK BR n BERG AAR 6 RK A SN ah AS AWA RS Heiss AAW Nrat® We inap uARA n+ BS BHB+2¥ 9) 1 fer AK FE WS shaw) Sh ha?” RR OMAR” eal BEAR” Se Ratt ane G Ran st 32 ES \ SS Cade Ht Ae IR AD M—jQahaey frag uw ARO ER ey B+ BE ORE mS IRIE OT FRR ENR a A RN ig mae AX By Wrarehy CaP Wath 4s A Ad 6 BE ae NBS Bom Be ke FA) saa 9 fined IG UR MEN IED iy ARTI RL A a Ral 1) BE KDA T Had ek Kemah» SBS al BES A 0 eh ws RIE RAS FD BEA WIN aR) WA BRE Ne Kam HAR TR ahh She No.8 y MAUR HOA WANAEN HEARNE :ete to the author without delay. It is apparent that at every step, from the receipt of copy by the editor to the actual run- ning of the press, the slightest error, inconsistency, lack of clearness in copy,or correction on author’s proof, causes delay, loss of time, extra and unnecessary handling of a mass of heavy metal, and also involves considerable expense. The old method of relying on the compositor properly to punctuate, paragraph, and correctly spell difficult scientific and foreign words in a carelessly pen- or pencil-written manuscript, and depending on the proof reader to detect and rectify errors of construction and grammar, has been abandoned. Publishers and print- ers have found the old method to be expensive and unsatisfactory, as compositors and proof readers are not properly equipped for this work. The complete codperation of author, editor, publisher and printer results in tangible benefits to all four. The author secures accurate. timely and creditable presentation of his ideas, with the irksome labor of proof reading and revision made as light as possible. The editor controls and is in touch with every step in the process of production. The publisher is enabled to predetermine results and obtains regularly and promptly the highest quality and best service for the amount expended. The printer makes a reasonable and assured profit by the economy of time and effort, minimum waste of material and the even distribution of work through the shop. THE LYMPHATICS SYSTEM OF THE GUINEA-PIG GEORGE K. HASHIBA From the Division of ‘Anatomy of the Stanford Medical School NINETEEN FIGURES INTRODUCTION This investigation of the lymphatic system of the adult male guinea-pig was entirely macroscopic. Owing to the technical difficulties the lymphatics of certain organs of the body such as the stomach were not injected successfully. Yet, in the more general sense, apart from the minor details, the fundamental plan of the system of the guinea-pig was revealed. The literature on the anatomy of this animal is very limited but the anatomy of the bones and the muscles by Alezais, ’98— ’02 has materially assisted me in this study. The method of investigation was as follows: The animal was killed with ether and while it was still warm, injections were made by means of the ordinary hypodermic syringe provided with a fine glass cannula. The animal injected was then care- fully dissected in the fresh or after temporary preservation. The needle must be of very fine caliber, indeed, in order to obtain a successful injection of the lymphatics of the guinea pig. The finest hypodermic needles obtainable in the market are too large, for their use usually causes a large area of the extravasation without filling any lymphatic vessels. Such a needle was, however, improved by fusing on to it a fine glass cannula, the tip of which was drawn out into a short but very delicate capillary tube. Among the various injection masses that were tested, Gerota’s Prussian blue and diluted India ink gave the most satisfactory results. 331 THE ANATOMIGAL RECORD, VOL. 12, No 3 APRIL, 1917 332 GEORGE K. HASHIBA This investigation is based upon experiments on about 30 male guinea-pigs, which after the injection, were carefully dis- sected with the aid of a watch-maker’s lens. Injection of the cutaneous lymphatics offered no little diffi- culty when I attempted to work from the outer surface of the skin, for the needle either penetrated too deep or too super- ficially. This difficulty was overcome by reflecting a small flap of skin by a crescentric incision and inserting the needle from the under surface. However injection must be made very slowly and under low pressure. Working with the mesentric nodes, it was found that when any of them were forcibly distended with injection fluid in order to inject the lymphatics backward to the loop of the intes- tine, the veins received retrograde injection instead as mentioned previously by Meyer, ’14. For the purpose of description the body is divided into the following regions: the head and neck, the upper extremity, the lower extremity, the thorax and the abdomen. The lymph- glands of the entire system are first described with reference to their average size, general shape, location and their afferent and efferent vessels. This is followed by the consideration of the lymphatic vessels of each region. The names of the glands are adopted, as far as possible, from the corresponding glands of the human lymphatics, but there are a few glands, which because of their closer similarity to those of cattle, are named after the work of Baum, 712. However, since some glands are situated quite differently in the guinea-pig from corresponding glands in man and other animals, it seemed better to adopt new names than to rigidly adhere to a given nomenclature. These glands were the following: A. Infra-mandibular node. From the efferent and afferent vessels, the node corresponds to the submental node. It les behind the mandible and the term inframandibular node seems quite appropriate. B. Dorsal and ventral pre-scapular node. These nodes corre- spond to the deep cervical nodes but since they lie at the level of the cephalic border of the scapula the term pre-scamies seems the more appropriate. — = LYMPHATICS SYSTEM OF THE GUINEA-PIG 333 C. Retro-scapular node. This belongs to the group of axillary uodes but lies wholly outside of the axillary space so that the term axillary node could only express a rough correspondence. Since it is more closely in relation with the caudal border of the scapula, the term retro-scapular seemed quite justifiable. D. Abdomino-inguinal and inguinal nodes. These nodes corre- spond to superficial and deep inguinal nodes respectively but their relation is so disturbed by the peculiar position of the hind limb that what might be the superficial inguinal node is pushed out of place and lies on the abdominal wall under cover of the tensor fascia lata, so that the term abdomino-inguinal is sug- gestive both of position and correspondence. The term inguinal node instead of deep inguinal is used because there is no superficial inguinal node. Consequently the adjectives ‘superficial and deep’ are superfluous and the term inguinal is employed to indicate a gland homologous with the deep inguinal node.! 1. THE LYMPH GLANDS Lymphoglandula parotidea (figs. 1, 15.) Position and size. The gland is found caudal to the base of the auricle, on the dorso-caudal border of masseter muscle and on the external maxillary vein. It is covered by the cephalic portion of the platysma and skin and is im- bedded in the parotid gland. Unless injected, it is very difficult to recognize it. It is flat and ovoid in shape, measuring 4 by 3 by 2 mm. Afferent vessels. The chief afferents of the auricle terminate in this node. Efferent vessels. One or two vessels from the node follow the external maxillary vein into the substance of the parotid gland and terminate in the submaxillary node which is placed at the junction of the external and internal maxillary vein and covered partly by the submaxillary salivary gland. Lymphoglandula inframandibularis (figs. 1, 2) This gland is located on the ventral surface of the anterior belly of the digastric muscle in close proximity to that on the opposite side. These nodes are usually placed transversely and one of them may be found quite behind (caudal to) the other. They are covered by the platysma and the skin. They are round and flat, 5 to 6 mm. in diameter and 2 mm. in thickness. 1 A supernumerary adrenal body was often found near the renal vessels. This was identified histologically. 334 GEORGE K. HASHIBA It is not, however, infrequent to find one of them considerably larger and more elongated and measuring 9 mm. in length. Afferent vessels. The lymphatics of the lower lip and those of the anterior two-thirds of the tongue terminate in this node. Efferent vessels. One or two vessels pass transversely between the sterno-hyoid and digastric muscles where they join with the collectors from the posterior part of the tongue. These conjoined vessels are directed caudal on the superficial surfaces of the sterno-hyoid and scalenus anterior and terminate in the deep cervical glands. Lymphoglandula submazillaris (figs. 1, 2, 15) Situation and size. This node which is entirely hidden by the parotid and submaxillary Lol. Farctidea Lymph Vessel to Sub-maxillary node ; pen StHtasseter ~~ = \ | / JCoplenius StTrapezius 7 St Auricularis Anterior oe * 6 hg Bub maxillarcs S1Orbicularis Oeuli BA » ; “ € S S _- Prescap: ularis a = a dorsalis Sn SAS ~ N.Omot Saris St Levator—__ _ __ a, = = Gl. RB see == = l Pre. ulahis MNasdabcalis — ventralis =< SSternocleédo \ mastoideus XN Bo ss \\ Lymph Vessels to yoin St Digastricus een if \ the thoracic dest Wa 7 | } | Lol Cerviealis The l. Side, and the Lgl Inframandikdarisy Le Oe Tight” one To The fugulo~ Cea aoe Sikelavian anglé Fig. 1 Superficial lymphatics of the neck /Sterno-cleide mavoideus 1 1 Ome-transVersarius Z 7 fis; ipleni us an / Pd | UnterTransversarias / A / MAngula iS Vs g/ SubmaxiMlaris — \ MMasseter \ Nigas Incus =~ > Lol. Prescapularisdarsalis Pas at ventralis ue s \ \ . %, i >< ; Ve \ \ \ \ MScalenus posticus Z gf. ‘nfra mandibularis” \ 1 \ \L gf .CerviealiS profunda \ Collectr prom the tongue ; I) Scalenus anterior S1Sternoh yo id Fig. 2. Deep lymphatics of the neck LYMPHATICS SYSTEM OF THE GUINEA-PIG 335 salivary glands lies at the junction of external and internal maxillary veins and is covered by the platysma. It is an irregularly shaped gland measuring about 4 mm. in diameter and 2 mm. in thickness. It is often represented by two glands; one medial to the other; but in very close relation to each other and separated only by the small por- tion of the submaxillary salivary gland and a part of the veins. When double these glands are connected with a number of short anastomos- ing lymph vessels. Afferent vessels. The vessels from a small portion of the auricle, from the frontal cutaneous area around the eye-lids, and efferents of the parotid node terminate in this node. Efferent vessels. Two or three vessels from this node follow the external jugular vein. At the lower border of the sternocleidomas- toids, the vessels take three different directions. One passes over the dorsal and the other over the ventral and dorsal prescapular nodes, Another vessel passes directly into the deep cervical nodes. Lymphoglandula prescapularis dorsalis (figs. 1, 2, 15). This node lies on the angularis muscle cephalad to the scapula and is covered by the omotransversarius and trapesius muscles. It is ovoid in shape, measuring 5 to 7 mm. long, 4 mm. wide and 3 mm. thick. Practically all the cutaneous lymphatics of the neck and those from the skin covering the cephalic portion of the scapula and the efferents from the parotid and submaxillary nodes terminate in this node. Two or three vessels are directed ventrally to end in the ventral prescapular node. Lymphoglandula prescapularis ventralis (figs. 1, 2). This gland is rarely absent. It lies at the root of the neck on the scalenus medius, and is covered by the clavicular portion of the sternocleidomastoid and platysma muscles. It is round and flat, measuring 4 mm. in diameter and 2 mm. in thickness. Two or three vessels from the dorsal prescapular and one or two from the submaxillary nodes terminate in this node. Usually two vessels pass ventro-cephalad under the sternocleido- mastoid muscles to the deep cervical nodes. Lymphoglandula cervicalis profunda (figs. 1, 2). This gland lies at _ the bottom of the V-shaped space formed by the sternocleidomastoid laterally, and the sternohyoid medially and is in contact with the trachea and anterior scalenus muscles lying in close relation to the internal jugular vein and the carotid artery. It is the largest node in the region of the head and neck. It is ovoid in shape and flattened, being 8 to 11 mm. long, 6 mm. wide and 3 mm. thick. All lymphatics from the head and neck finally terminate in this node. It receives the collectors from the tongue and the efferent ee from the submandibular, submaxillary and ventral prescapular nodes. The efferent vessel from this node is the largest lymph vessel in the neck. It is 12 to 15 mm. long, distinctly sacculated and closely fol- lows the internal jugular vein, lying between the sternocleidomastoid 336 GEORGE K. HASHIBA and the anterior scalenus muscles. At the root of the neck it ter- minates on the left side at the cervical bend of the thoracic duct and on the right side, at the junction of the internal and external jugular veins. 2. The lymphglands of the anterior extremity Lymphoglandula retro-scapularis (figs. 4, 5). This node is located on the outer surface of the latissimus dorsi at the angle formed by the caudal border of infraspinatus and the dorsal border of the triceps brachialis. It is imbedded in a pad of fat and covered by the pannicu- lus earnosus and the skin. It is ovoid in shape and flat, 9 mm. long, 4 mm. wide and 2 mm. thick. M Retoralis MM Cotoco-bachialis i! y, ~MCleidomastideus Tendon of M. Biceps /1Subclavius ve \ \ se be ei a Sa angle ae Tol Pailvis Fh /MDelto-Clavicularis es Axilbris Pima Costae /1Slerno-costalis /7. Bice; S g /1Scalenus Pot Lal. A ywihlaris Si 7 Reclus Abaominis /1 Peetoralis IM Trceps(medial head) /MLatissrmusDorsi MSerratus SI dntercosialis Anterior Uh Toros lI] poe apularis Fig. 3 Lymphaties of the axilla The lymphatics of the outer surface of the anterior limb, and the dorsal and lateral cutaneous lymphaties of the thorax terminate in this node. Two or five vessels from this gland penetrate the latissimus dorsi and terminate in the lymphoglandula axillaris. Lymphoglandula axillaris (figs. 3, 17). This gland lies on the medial surface of the teres major at the angle formed by the axillary artery and the tendinous insertion of the latissimus dorsi. Medially it is in rela- tion with the scalenus anterior. It is a cireular node 5 to 7 mm. in diameter and 2 mm. thick. It receives efferent vessels from the retro-scapular node and the inner cutaneous lymphatics from the pectoral region also terminate in it. The large single trunk which follows the axillary artery is imter- rupted by the lymphoglandula axillaris prima costae. However, it is not infrequent to find this trunk dividing near the latter node into two LYMPHATICS SYSTEM OF THE GUINEA-PIG (1 Sta pulo-Clavetabis posterior Pha hosimusDors, {pene vessel To Ebi Arille us [Dette Acromia /iwcepatteny head) WM Delle-Chnedlans MN pannel Carnosus ABrach/al is Shaeps (Lateral head) [Flexor Canpthnarts 41. Biceps Uhshanse? Carpe Unaris TM Extensor Carp, Kakelis 17 Extensor Mimiom Prigihe \ (Mislensor Tehers (1 fansor Comm Digi Fig. 4 Lymphatics of the fore limb Crest z Mium MObligus Abdominis ; ; ay /1 Glulexas Ped extlernus Hip 49l.abdomino- MGlutews Mim 4ngutnalis Great Sacrasciatic Foremen St Tensorfasecae 11P, riformis Lalae JA Scho-coccygeus NIschiadious MQnadriceps MObluratarinternus M Glulews max MNGemell bg A = 2A A y% ro “ iB MQuadratus Femoris \- = i Kh ‘ : BZ (f \ . Cz Biceps Ny ‘ /1Semi-tendéinosus \ Lol. Poplilea “1 Gastroonemias Fig. 5 Lymphaties of the thigh 337 338 GEORGE K. HASHIBA vessels, one of which terminates in this node and the other at the junction of the external jugular and subclavian veins. Lymphoglandula axillaris prima costae (figs. 3, 17). This node is found on the outer surface of the first rib, at the termination of the axillary blood vessels, being covered by the pectoral muscles. It is circular and about 5 mm. in diameter and 2 mm. in thickness. alee The efferent vessels of the lymphoglandula retro-scapularis and very often some efferents of the lymphoglandulae tracheobroncheales and lymphoglandula axillaris end in this node. A large single trunk leaves the node and terminates at the junction of the subclavian and external jugular veins. 3. Lymphatics of the lower extremity Lymphoglandula poplitea (figs. 5, 7, 15). The node lies imbedded in the popliteal fat, in the triangular space bounded by the biceps femoris, semitendinosus and gastrocnemius muscles. It is spherical in form and about 2 to 3 mm. in diameter. The lymphatics of the lateral surface and dorsum of the foot termi- nate in it. The majority of the efferent vessels follow the popliteal blood vessels where they join with the deep lymphatic vessels of the hind: limb. However, there is one other efferent vessel which passes along the posterioro-caudal border of biceps the femoris. Near the origin of the latter it crosses the outer surface of the muscle and soon passes under the gluteus maximus to accompany the ischiadie nerve with which it enters the pelvic cavity through the foramen ischiadicum majus. In the pelvis it lies close to the median line on the anterior surface of sacrum and finally terminates in the hypograstric node. Lymphoglandulae abdomino-inguinales (figs. 5, 6, 7, 15). These glands are found along the course of the superficial circumflex ilac vessels between the obliquus abdominis externus and tensor fascia lata muscle. This group consists of 2 to 4 nodes of varying size, the largest of which reaches about 7 mm. in diameter. All cutaneous lymphatics of the abdomen and the hip, the lym- phaties of the outer and inner surfaces of the thigh, those from the scrotum and from a small portion of the lateral part of the hind leg terminate in this node. The majority of the efferent vessels accompany the superficial cireum- flex iliac vessels and terminate in the inguinal nodes but a small vessel passes through the femoral ring to end in the iliae node. Lymphoglandulae inguinales ( figs. 6, 7). This pair of nodes is found on the femoral vessels, close to the femoral opening and is composed of a medial and a lateral node. The medial node is irregular in shape measuring about 5 mm. in diameter and 2 mm. in thickness. The lateral node is the smaller of the two. It is circular and measures about 3 to 4 mm. in length and 1 mm. in thickness. LYMPHATICS SYSTEM OF THE GUINEA-PIG egl.h ipegastriea iliaeas efferent «¢ i Popliteal node. (1 Pseas a ee Lgl. abdemine - inguinales 4 11. fensor fasciae < S Se M7. Quadtafus femoris 1 Rtetus intornas MS anferior 1 hee/us tuhepmas Pos ter/op Salel; pr of Saphenous Vern ‘9, mgue males WA pyri form's mM. obf/urator imternus (gl. jaea wxtirna, Fig. 6 Lymphaties of the femoral and pelvic regions Lo). Hypogastrica -—__ ' ares = MM Llio- psoas Milio-coceygeus S-Se 2 MobtursiorInternus — —~ lh ‘ —— ~-Lol Lliace-Externa Jz ¢ a Stig > i wo - - At7ensarfesciaeLatse g —— Lp. Inguinales MReclus InfernusAnterjor - -- MPuadricepsfemerrs MAddefor-medius ——-~ x , io ~M Fectmeus — -M Add Nit Z lector magmas pea Cart ¢ Cord 4 ! | A Blae ; c/3 oe L Poplifea M.Semimembranosus — — ; 49 ‘op + —S1Gatreenemius / ~~-M.Semitend inosus Fig. 7 Lymphatics of the abdominal and femoral regions 339 340 GEORGE K. HASHIBA The efferents from the popliteal nodes, the deep lymphatics of the hind limb and those accompanying the saphenous artery terminate in the medial, while the efferent vessels of the abdomino-inguinal nodes terminate in the lateral node. These two nodes are in communication with each other. A number of vessels pass through the inguinal canal and terminate either in the external iliac or in the common iliac nodes. 4. The Lymph glands of the abdomen Lymphoglandula iliaca externa (figs. 6, 7). This is a very inconstant gland located when present on the external iliac vessels. Ii is quite round and 3 mm. in diameter and 2 mm. in thickness. It is inter- polated in the course of the efferent vessels coming from the inguinal nodes but sometimes also receives afferents from the bladder. Lymphoglandula hypogastrica (figs. 6, 7, 10, 16) This is a small but constant gland lying in the pelvie cavity at the origin of the hypo- gastric artery. It is a small circular gland 2 mm. in diameter and 1 mm. in thickness. The lymphatics of the bladder and the seminal vesicle and the efferent vessel from the popliteal node terminate in it. An inconstant number of efferents cross the common iliac vessels to terminate in the common iliac node while certain other vessels pass toward the abdominal aorta, contributing to the formation of the lymphatic plexus around this vessel and the inferior vena cava. Lymphoglandula iliaca communis (figs. 6, 7, 10, 16). This node is placed at the angle of abdominal aorta and the common iliac artery. Indeed it is not infrequent to find it lying entirely on the outer side of the common iliac vessels. It is fusiform in shape, 7 mm. long, 3 mm. wide and 2 mm. thick. The entire lymph stream of the lower extremity and from the pelvic cavity reaches it. Vessels from the bladder and from the pelvis to- gether with the efferent vessels of the inguinal, the external iliae and the hypogastric nodes terminate here. As soon as the efferent vessels from this node reach the side of the great abdominal vessels, they form a very rich plexus which entirely surrounds these vessels. Their final destination is the cisterna chyh, but before reaching it they are interrupted by small ‘Schaltdriisen’ scattered in the course of the plexus. Lymphoglandulae mesentericae intestinales (fig. 9). A large number of lymph nodes of varying size are found between the two layers of the mesentery. Among these is one large node which lies on the main trunk of the mesenteric vessels. This node receives the entire lymph stream of the intestine. The rest of the nodes are scattered on the peripheral branches of the same vessels, constituting glands for differ- ent segments of the intestine. These latter glands are quite variable. Hence the description of a single type of arrangement can not be entirely satisfactory. Nevertheless, there is a certain prevailing ar- rangement which is considered here as typical. —— OO —— a LYMPHATICS SYSTEM OF THE GUINEA-PIG 341 The following glands constitute the lymphoglandulae mesentericae intestinales: . . The lymphoglandula colicae descendentis. . The lymphoglandula colicae transversalis. . The lymphoglandula colicae ascendentis. . The lymphoglandula intestini tenuis. Lymphoglandula ilio-coecalis. Lymphoglandula coecalis. Lymphoglandula mesentericae communis. Be NOOUPRWNHE . Lymphoglandula colicae descendentis. This node is found close to the middle of the descending colon, imbedded in a small quantity of fat. It is somewhat round in shape, 2 mm. in diameter and 1 mm. thick. Lymph from the descending colon drains into this node. Lol Tracheo-bronchialis Arch of aorta P = Sf LAS , Left autteulo-ventricular A.Palmonis yaa = . of ps al Right auricle ; Lett auricle at IPO Fight. Quijceulo-Ventricular Vena ped: 7 collector Lal. Inter-bronchialis Fasterior interventic ular Collector Vena Cava inferior Vena Pulmonis Fig. 8 Lymphatics of the heart One small vessel from it follows the inferior mesenteric artery and at the origin of the latter joins with the lymph plexus surrounding the abdominal aorta. 2. The lymphoglandula colicae transversalis is located close to the middle of the transverse colon. It is round in shape, 5 mm. in diameter and 2 mm. in thickness. The lymphatics of the transverse colon end in it and the efferent vessels usually reach the common mesenteric node. 3. Lymphoglandula colicae ascendentis. This is found close to where the ascending joins the transverse colon. It is 4 mm. long, 3 mm. wide, 3 mm. thick and is not rarely accompanied by other very small glands close to it. The majority of the afferent vessels draining into this node are from the ascending colon but a few vessels from the transverse colon also end here. Lymph from it is received into the common mesenteric nodes. 342 GEORGE K. HASHIBA £ gl. ColicaeTrans versalis Pincreas¥ Duodenum Ar. Superior Pnesenlerica Tortal Vein % Lymphatic Trunke to cisferma chyli L Leer = Py Lgl. Tntest tenuis Lol Mensore pe ye; ; Lal Coela /i$ L_ Lal Colic. Lgl Aoriae - U a abdominalts y of aren centee ant ¥ Lymph Vessel Ascending Colon Abdominal aorla © Inferror Vena Cava Colon. Lal Tiiocoecal. SE Fig. 9 Lymphatics of the intestine The hypogastric Node recelving Lymph AT frm seminal Vesicle { MING Y cara wz arlerior part of \ | \\ Wy / cai Bladder. fhe gl Miacae Communis Lymp vessels. recetving ts yiph Seminal vesicle ‘rom posferror pat of Bladder later « fee pens i hve te Reetuslniernus /1Sphinclet a ! i (\ rt —VasDeferens ima y i Urethrae membronaceae yi, Lymphatic nef-wark Sg Glans Fig. 10 Lymphatics of the genito-urinary organs LYMPHATICS SYSTEM OF THE GUINEA-PIG 343 4. Lymphoglandula intestini tenuis. The gland of the small intes- tine lies near the center of the mesentery of the small intestine. It is an irregularly shaped node about 10 mm. in diameter and 2 to 3 mm. in thickness. A large number of afferents from the small intestine converge toward it. Two or three vessels from this gland terminate in the common mesenteric glands. 5. Lymphoglandula ilio-coecalis. A glandular mass constituting the ilio-coecal node is located around the spot where theileum and coecum _meet. This composite mass may be grouped into onenode measuring about 10 mm. in diameter and 4 mm. in thickness but it is more often divided into two unequal masses. The lymph vessels from a small portion of the ascending colon and from the caudal half of the coecum terminate in it. Two or three vessels from this node traverse the mesentery stretched between the coecum and ileum to reach the coecal node. 6. Lymphoglandula coecalis. This node lies in the mesentery be- tween the ileum and the coecum close to the common mesenteric node and the gland of the small intestine. Sometimes it fuses with the gland of the small intestine. It is somewhat elongated, measuring 7 to 10 mm. long, 3 mm. wide and 2 mm. thick. Many of the coecal lymphatics terminate in this node, as do also the efferents of the ilio-coecal node. ‘he or three large vessels from it end in the common mesenteric node. 7. Lymphoglandula mesentericae communis. This is the most im- portant gland of the intestine. It lies on the trunks of the mesenteric blood vessels just before these cross the duodenum and is in close rela- tion with the lymph gland which drains the small intestine and with the transverse colon and the head of the pancreas. It is often divided by the pancreas, one portion lying anterior and the other posterior to it. Behind this node lies the apex of the coecum and a portion of the duodenum. It is irregular in shape, about 8 mm. long and 3 mm. thick. In a number of cases it is not a separate node but forms one large mass with the glands of the small intestine and the appendix. However, separation into three distinct nodes is more common. With the exception of the lymphatics from the descending colon, the entire lymph from the intestine drains into this node. Its afferent vessels come from the coecal, ilio-coecak and intestinal nodes. The single large distinctly sacculated truncus lymphaticus intestinalis which leaves this node closely follows the course of the superior mesen- teric artery and empties into the cisterna chyli. Often a small vessel is sent from this trunk to the retropancreatic node. Lymphoglandula retropancreatica (fig. 11). This gland is found behind the head of the pancreas on the superior mesenteric vein near the foramen epiploicum. It is a pear-shaped or somewhat ovoid node about 8 mm. long. 344 GEORGE K. HASHIBA A number of vessels from the liver and a small vessel from the com- mon mesenteric node end in it. One to four vessels are given off from this node and either join with the truncus intestinalis or follow the superior mesenteric artery and terminate in the cisterna chyli. Lymphoglandulae aortae abdominalis (fig. 16) There are a large number of glands around the abdominal aorta and the inferior vena cava. These glands are divided into three groups: the inferior, middle and the superior. But it must be remarked that these divisions are absolutely artificial, for the nodes form a continuous chain without any distinct separation. Hence it is difficult to group separately. Some lymphatie vessels go to certain of these nodes but the territory drained is not distinct. Liver-lobes ) Ga// Bladder Gastro hepatic + Mepato-duodena/ Foramen - Epiploicum j SSS Kidney 7 y bs 4 Li Ntrmeverse Colon / ‘ j i) Infetior Vena Cava’ 4 ph oer Mead of Pancreas Lgl Retroparcreatica/ fal leserterica around which J _ Commuris ) , Ferio! vein Ouodenum /oops Fig. 11 Lymphaties of the liver 1. The lymphoglandulae aortae abdominalis inferiores are composed df three to six glands about the size of a pin’s head which are grouped around the lower half of the great abdominal vessels, in meshes of the lymphatic plexus. These nodes are nothing more than ‘Schaltdriisen’ interrupting the lymph from the hypogastric and the common iliac nodes and from the descending colon on its way to the cisterna chyli. 2. Lymphoglandulae aortae,abdominalis mediales. This group of nodes is found around the abdominal aorta, the renal vessels and the superior mesenteric and spermatic arteries. The glands of this group are much larger than those of the inferior mesenteric arteries. They are usually ovoid in shape and the largest one may be as big as 5 mm. but very many small nodes may be found among them. Since these glands lie in a plexus of lymphatic vessels, some of them are only interrupting nodes while others receive definite afferents from the kidneys and the testicles. LYMPHATICS SYSTEM OF THE GUINEA-PIG 345 The efferent vessels from them terminate in the cisterna chyli which lies behind the aorta, between the pillars of the diaphragm. 3. Lymphoglandula aortae abdominalis superior. This node lies against the lateral surface of the pillars of the diaphragm. It is ovoid or round measuring about 4 mm. across. . aS Testicles ——, Fig. 12 Lymphaties of the testicle and kidney Manubrium Taferruph: ng yode V2 TAN Lretitic mesa N e RSA Mi OK A y \ WK y NS se oY. oe iy ‘Diaphragm Gi fps gn ~ SS ZZ Fig. 13 Lymphaties of the diaphragm 346 GEORGE K. HASHIBA Some afferents from the kidney and others from the abdominal plexus, which do not empty into the cisterna chyli at this level join it. Usually one of these vessels either penetrates the pillar of the diaphragm or passes behind the arcuate ligament and joins the cis- terna chyli at once. 5. The lymph glands of the thorax Lymphoglandulae arteriae mammariae internae (figs. 13, 17). One node is located internal to the first rib in company with the internal mammary vessels. Cranially it is in relation with the origin of the sterno-hyoid and sterno-thyroid muscles and dorsally it is in relation with the innominate vein and the arch of the aorta. It is a small, somewhat round gland, measuring about 3 mm. in diameter. Cesppegn Lol Trachec-hronchialis Lollnter-bronchialis: Oeso us Fig. 14 Lymphaties of the lung Lymphaties from the diaphragm and from the intercostal spaces terminate in this gland and it is usual to find that some vessels from the tracheo-bronchial nodes join it also. The efferent vessels from the nodes of both sides form complicated plexuses on the innominate vein. The plexus on the left side joins with the ductus thoracicus and on the right with the cervical and the subclavian lymph trunks just before their termination in the vein. Lymphoglandula inter-bronchialis (fig. 14). A single node is located at the bifurcation of the trachea between the roots of the bronchi. Ventrally it is in relation with the pulmonary vein and dorsally with the oesophagus. It is a somewhat spherical node measuring about 3 mm. in diameter. LYMPHATICS SYSTEM OF THE GUINEA-PIG 347 ‘ The lymphatics of the lung and heart send some of their lymph ves- sels to it and the efferent vessels terminate in the tracheo-bronchial nodes of both sides. Lymphoglandula tracheo-bronchiales (figs. 14, 17). These nodes lie in the angles between the trachea and bronchi on the right and left side respectively. They are elongated thick nodes 10 mm. long, 4 mm. wide and 3 mm. thick and are often separated only by the azygos vein which crosses the united nodes. They receive afferents from the lung and the efferents from the bronchial node. The lymph vessels joining the glands of both sides of the trachea are often found to anastomose with each other and also with a number of anastomatic vessels which surround the trachea. The efferent vessels from this node terminate in three possible places: in nodes along the internal mammary artery, in the lympho- glandulae axillaris prima costae or in the plexus in front of the innomi- nate vein. Il. THE LYMPH VESSELS 6. The lymphatic vessels of the head and neck 1. The lymphatics of the ear (figs. 1, 15). The lymphatics of the ear are best injected on the medial side. The network appears near the margin and from this plexus appear a number of vessels which join with each other as they approach the root of the ear. The majority of these radicles pass to the parotid node but a few pass directly to the submaxillary lymph node. 2. The lymphatics of the face (figs. 1, 15). A large number of very fine vessels arise in the frontal region from the eye-lids, the side of the nose and from the upper lip. They all run toward the lateral sur- face of the neck, penetrate the platysma and come to lie between it and the masseter muscles. The larger collectors, however, follow the in- ternal maxillary and branches of the external maxillary vein. All these vessels converge to the submaxillary lymph nodes. The lymphatics of the lower lip terminate in the inframandibular node. 8. The cutaneous lymphatics of the neck (fig. 15). All the cutaneous lymphatics of the neck which are subcutaneous at first penetrate the platysma. They then pass either through the dorsal or ventral border of the omotransversarius and terminate in the dorsal prescapular node. 4. The lymphatics of the fore-limb (figs. 3, 4, 15). When injections are made into the pad of the foot of the anterior limb, several vessels are seen to pass centrally in various directions. Some pass between the digits, others curve around the radial and ulnar borders of the foot to appear on the dorsum and still others pass to the flexor surface of the forearm. The latter have an entirely different path and termina- tion from the former. Those that appear on the dorsum of the foot pass to the extensor surface of the forearm where they join to form one THE ANATOMICAL RECORD, VOL. 12, No. 3. HASHIBA GEORGE K. 348 soryeydurA] snooueynod ayy, ey ‘Ay Sn m24D4SPD HY He SyroIp PY RID Lo8u%yXF bh, EN OMB BRAUN Lf, f ' vushpojd fo phe 7 doag- eS : Mel XZ) | XY Wik ae / Pe ot oe an f i 7 : a) aromas ita anne 4 HERE matetp] tases Ww, \ | \ \ y SHOY/xDMEgn yor Se ) / eh ty / \ / SIUOU If) LID 1084 94XF yf, ees Wi neo y a Wy é % ‘a = / / / 3 \ i Suu) 11109 40X2/'f py ha oprapeo cess // / 7 SPOR Ly \ iy s smog 7 / Fo i) / Poe rum OuO Ly / ihe / Sabi 177 7° SIfOrTepowrny is dozene zy, / - he ee PON (PppeL: - Y /, WY Yi 4e sdaoigyy / Z ; yp t- m 4 SL yrn0 sro / “BLE ropld 00504 be A pe oh a / 1 Suouaysnpoeyy 7! Ret OM a /°7 PN ‘. worysobodhy by / / 1 ‘ jor Sermanoenel joy eel | W hiyge, Li ‘\ SUD/M2 Ung I0.UaI Ee y/ i: UE u ' ; / reoqisnumsssyo7 ty jf 1 PSSA luk fi LA es OU ; uDroung ibe LLY F SEFOLOUPOS CLOSELY =! i If fe ai bp \ i iy, wohyysartyy 1 Peropyue. | * : SMX SPAN ! / / RR as if 6 snsousroy snjno}uurey f] dor, py fi coushyoyy jy! LYMPHATICS SYSTEM OF THE GUINEA-PIG 349 vessel which accompanies the cephalic vein to the middle of the bra- chium. Here it leaves the vein and penetrates the panniculus carnosus, terminating at once in the retroscapular node. Those vessels which appear on the flexor surface of the forearm con- tinue their course as far as the distal third of the brachium where they meet the brachial artery. Here they join with the deep lymphatic vessels which accompany this artery and end in the axillary node. 5. The lymphatics of the hind limb (figs. 5, 6, 7, 15). A number of lymphatic vessels which arise from the medial and lateral margins of the foot arrange themselves on the lower part of the leg into medial and lateral collectors. The medial collector follows the saphenous vein very closely and terminates in the inguinal nodes. The lateral collectors pass along the outer surface of the leg and the majority terminate in the popliteal node. However, one or two vessels continue their course over the thigh and end in the abdomino-inguinal node. Indeed, the entire cutaneous lymphatics of the thigh converge to the latter nodes. The deep lymphatics of the hind limb were not successfully injected except those along the femoral artery in the region of the thigh. If the pressure is relieved along the femoral artery by cutting away most of the adductor muscles, injections into the popliteal node always fill the deep lymphatic vessels which form a plexus around the artery and terminate in the deep inguinal nodes. 7. The lymphatics of the abdomen 1. Cutaneous lymphatics of the abdomen (fig. 15). Fine lymphatic vessels which arise from the skin of the abdominal wall soon penetrate the panniculus carnosus and join with one another as they converge to the abdomino-inguinal nodes. Separation of this area from that of the thorax is not sharply marked off for one merges into the other. Hence, an injection in the wide common zone which constitutes the boundary between them, may be seen to travel either to the external retroscapular or to the abdomino-inguinal node. 2. The cutaneous lymphatics of the perineo-pudendal area (fig 6). From the network of origin on the skin in the perineal and pudendal regions arise one or two collectors which pass between the thigh and the abdomen to end in the abdomino-inguinal node. 3. The lymphatics of the bladder (fig. 10). A fine lymphatic network from which a number of collectors leave is found on the bladder. These collectors are divided into anterior and posterior collecting trunks from the anterior and posterior surfaces of the bladderrespectively. The anterior gain the brim of the pelvis and usually joi the common iliac nodes while the posterior collectors run to the floor of the pelvis to reach the hypogastric node. 4. The lymphatics of the seminal vesicles (fig. 10). From the very fine network of origin on the surface of the seminal vesicle arise a number of collectors which join with each other at the free border of 350 GEORGE K. HASHIBA the mesentery of the vesicle to form one common vessel which at the side of the bladder joins with the collectors from the posterior surface of the latter. This conjoined vessel runs over the floor of the pelvis to reach the hypogastric node. 5. The lymphatics of the penis (fig. 10). The lymphatics of the prepuce and glans form two collectors which run along the dorsum of the penis. They pass under the symphysis of the pubis, over the sphincter urethrae muscle. Close to the bladder they diverge and after reaching the pelvic brim follow the external iliac vessels and terminate in the common iliac nodes. 6. The lymphatics of the testicle (fig. 12). When injections are made into the substance of the testicle a large number of fine vessels appear on the surface. These vessels converge to the point where the sper- matic vein leaves the testicle where they form common collectors. These collectors, two or three in number, follow the spermatic vessels. The artery and vein are closely interwoven by these lymphatic vessels in their entire course. Near the kidney, the lymphatic vessels leave the artery, but follow the vein and after reaching the wall of the renal vein terminate in some of the nodes of the middle abdomino-aortic group. 7. The lymphatics of the kidney (fig. 12). The deep lymphaties of the kidney are easily injected by simply forcing the injection mass into the kidney substance. A number of lymphatic vessels appear imme- diately at the hilum. These surround the renal blood vessels and terminate in the nodes in the middle abdominal aortie group. 8. The lymphatics of the intestine (fig. 9). Working out the lym- phaties of the intestine is a difficult task in a small animal Owing to the delicacy of the intestinal wall, direct injection with the needle was unsuccessful. In the caecum and a small portion of the large intestine, however, injections made on the teniae were quite successful in demonstrating the fine plexus on the walls. To inject the lymphaties of the other portion of the intestine, the following method was employed: (1) the intestine was cut into pieces about 8 cm. long, the mesentery being kept intact as much as possible. The contents were washed out thoroughly and a small quantity of rather coarse sand was introduced. This was rubbed against the intestinal wall by rolling the intestine between the fingers, so as to injure the interior of the intestine. The intestine was then flushed again and every drop of water pressed out. Into a piece of the intestine so treated the injec- tion mass was then introduced quite fully and the ends clamped. By massaging gently with the fingers, the injection fluid was seen to slowly fill the lymphatics. The lymphaties of the small intestine were also very well shown by the historical method. An animal which had fasted for a day, was fed with milk. After about one hour it was killed and the intestine examined. This method showed the collectors beautifully as fine white lines which could be traced to the cisterna chyli but their network of origin in the wall of the intestine was not recognizable. LYMPHATICS SYSTEM OF THE GUINEA-PIG 351 The lymphatics of the intestine arise from a very closely meshed plexus in its wall. From this network several collectors run which pass through the glands placed in their course and finally form two efferents. One of them is the efferent of the node of the descending colon, which, following the inferior mesenteric artery contributes to the formation of the lymphatic plexus around the abdominal aorta. The other is the truncus intestinalis which connects the common mesenteric node with the cisterna chyli. This latter trunk carries the lymph stream from the entire intestine, except from the lower portion of the colon. 9. The lymphatics of the liver (fig. 11). The deep lymphatics are easily shown when injections are made into the hepatic tissue. A number of collectors appear at the hilum and pass together with the cystic duct and the superior mesenteric vein, between the two layers of the hepato-duodenal ligament in front of the epiploic foramen and reach the retropancreatic node. 10. The lymphatic plexus around the abdominal aorta and inferior vena cava (fig. 16). There is a very rich, closely-woven lymphatic plexus around the great abdominal vessels extending from the bifurcation of the abdominal aorta to the cisterna chyli. This plexus is further com- plicated by the presence of a number of glands, scattered in the meshes. These I classified as lymphoglandulae aortae abdominalis inferoris and medialis. The following are the contributors to the formation of this plexus: efferents from the common iliac and hypogastric nodes, those from the descending colon and the collecting vessels from the kidney and testicle. This plexus is finest in its lower course and gradually forms itself into larger trunks as it continues upwards. Near the renal vessels, behind the aorta, between the pillars of the diaphragm it becomes a large cisterna chyli, into which the truncus intestinalis empties. 8. The lymphatics of the thorax 1. The cutaneous lymphatics of the thorax (fig. 15). The ventral lymphatics cover the pectoral area. The collectors which at first are subcutaneous penetrate the panniculus carnosus and lie between it and the pectoral muscles. They join with each other as they pass toward the axilla and converge to the axillary node. The dorsal lymphatics cover the dorsal and lateral surfaces of the thorax and also the caudal portion of the scapular region. The col- lectors from this area, after penetrating the panniculus carnosus con- verge toward the retroscapular node. 2. The lymphatics of the diaphragm (fig. 13). The lymphatics of the diaphragm were injected only on the ventral surface of the muscular portion. A large number of the vessels are seen to join with one another on the diaphragm just behind the xiphoid cartilage whence they continue their course as the internal mammary trunks. If the injection is made on the diaphragm a little laterally, only a few ves- sels appear. These traverse a few lower intercostal spaces obliquely and likewise reach the internal mammary trunk. S52 GEORGE K. HASHIBA 3. The lymphatics of the intercostal spaces (fig. 13). The lymphatics of the intercostal spaces were injected successfully only in the anterior (ventral) portion. The vessel from each intercostal space has a slight cephalic obliquity. It traverses the intercostal space just beneath the parietal pleura and the transversus thoracis muscle and joims the the internal mammary trunk almost at right angles. VJugularis Externa q 4%; wis el 5 USubclavia ee f UInnomi eae ef: VSubclavia Ducts thora CICUS Cisterna Chyli Ve 9! Aortae abdominals Lq/ Aorta abdominalis Superior Superror Ar re Lg] Aortae a bdominalis BON oe medialis Truncus intestinalis AMesenterica Lgl Aorfae Lal Aortae abdominalis Aabdominalis medialis inferior py Ey¢eerent the glan We eA pi A.Mes enterica inferior ify Commun Lgl liacaeCommunis Lgl.t ae Allaca externa of, g/. Hypogastnica op aetesel Atypogastrica Fig. 16 The main central lymphatic nodes and thoracie duct LYMPHATICS SYSTEM OF THE GUINEA-PIG 353 4. The internal mammary trunk (fig. 13). The internal mammary trunk lies directly lateral and internal to the sternum accompanying the internal mammary vessels. It is in relation ventrally with the intercostal muscles and dorsally with the transversus thoracis and the parietal pleura. It receives the lymph from the intercostal spaces and the diaphragm. In about 60 per cent of the cases, this trunk is interrupted by a small node placed a little above the middle of its pee and behind the first rib. It opens into the internal mammary node. : 5. The lymphatics of the lung (fig. 14). The deep lymphatics of the lung can easily be demonstrated, for by injecting deep into the lung tissue a number of vessels appear at the hilum. These vessels, at first, 2 = VJugulare interna —— | eee a I Cervicalis ‘ . 4 rR f goes \ & 7 C] £ | 4 f —V.Jugularis extema : y \ E : f \ 4 : 4 Lol Axillars \ fi 9 J ‘Prima costae \ by: vd __VAnillaris eb ee 7, SAV ne ~~~Lo/ Axillavs WF f\\f “~~ Lol Mammaitae we >) Deeplessel 7. me. we intone WY) \9 A hg From Retrosca- — pular node ——Ductus Thotecseus Fig. 17 The terminal vessels surround the bronchi and then reach the tracheo-bronchial and bronchial nodes. 6. The lymphatics of the heart (fig. 8). The majority of the fine vessels which arise from the surface of the ventricle are directed toward the anterior and posterior interventricular groove and but compara- tively few vessels pass to the auriculo-ventricular grooves. Thus there result in the heart, four main collectors: the anterior and posterior interventricular and the right and left auriculo-ventricular collectors. The anterior interventricular collector follows the artery of the same name and a little above the middle of its course divides into two vessels which after receiving the auriculoventricular collector pass to opposite sides of the pulmonary artery, whence they run under the arch of the aorta, and terminate in tracheo-bronchial nodes on both sides of the trachea. 354 GEORGE K. HASHIBA Se = >= erly at e te: PEE =e, DOVES ae