The Identification of Acidophilic Cells in the Human Pars Distalis-'

JAMES L. CONKLIN Department of Anetonz?g, The University of Michigan, Ann Arbor, Michigan

ABSTRACT Sections of human hypophyses were fixed in either formalin or Bouin's fluid and stained by a variety of acid stains. The stains were employed singly, in combination with each other and in conjunction with various mucoid staining pro- cedures, After these procedures the chromophils of the pars distalis were classified as either acidophilic, mucoid, acidophilic-mucoid, modified or degranulated. The remain- ing cells which lacked specific chromophilic granules were classified as chromophobes. The acidophils after the terminology of Ezrin were designated as cell types I, I1 and VIII which correspond to the alpha, eta and epsilon cells of Romeis. Numerous, carminophilic type I cells were present in glands of all ages. Although usually pyramidal or oval in shape they also exhibited a variety of morphologic forms. The type I cells were further subdivided into small, light and dark cells on the basis of size and nuclear characteristics. The type I1 cell although present in all glands was most frequently observed in fetal and post-menopausal pituitaries. It was tinc- torially identified by the prominent staining of its cytoplasmic granules with erythrosin. The type VIII cell exhibited staining properties intermediate between cell types I and 11. On the basis of tinctorial and morphologic properties it was tentatively identified as a modified type I or type I1 cell.

Prior to a study of pituitary cytogenesis phologic parameters are variable and de- the literature was searched for previous pendent on the physiological state of the descriptions of pituitary cell types and the cells; (3) cells that secrete specific hor- methods utilized for their demonstration. mones that are chemically consistent can It was soon apparent from this literature be expected to exhibit consistent histo- survey that a great deal of confusion ex- chemical properties in spite of morpho- isted about the type and function of pitui- logic variability. tary cells and the specificity of staining In order to clarify the cytology of the methods employed (see reviews : Purves, human pars distalis,2 sections were ex- '61; Halmi, '63; Ezrin, '64; and Herlant, amined after the application of a variety '64). of histological and histochemical proce- Factors which apparently contributed to dures. The specificity of the methods was this confusion were : ( 1 ) the utilization of evaluated and the chromophils of the nor- selective but not specific staining methods mal human hypophysis were identified. to study glands from different species, (2) Additionally, the histochemical cytology of a lack of adequate morphological descrip- the gland was compared with the pub- tions of specific cell types, (3) a lack of lished immunocytologic descriptions. The consistency in the use of nomenclature present report deals with a description of and (4) overemphasis an the correlation acidophils and chromophobes, while a sub- between cell types and concomitant en- sequent report will describe the hypophy- docrine disorders. seal mucoid cells. For the sake of brevity Presumably, less confusion would have and in observance of the above criticisms, occurred if certain guidelines common to only limited reference is made to studies all cytological studies had been observed. of the hypophysis of other species.

Some of the more significant of these are: 1 Research supported by grant HD 00557 from Na- (1) cells of different species which have tional Institute of Child Health and Human Devcl- opment, National Institutes of Health. the same tinctorial properties may not nec- ZThe term pars distalis is employed as defined by Purves and Bassett ('63) and is homologous to the essarily have the same function; (2) mor- pars anterior and of most species.

ANAT. REC.,156: 347-360. 347 348 JAMES L. CONKLIN

MATERIALS AND METHODS differentiated in aniline alcohol (0.1 cm3 The material' employed in the study aniline oil: 100 cm' 95% alcohol) consisted of 2,500 3-5 cr sections of human (Romeis, '40). hypophyses obtained from 15 fetuses and 2. 2% orange G (C. I. 16230) in 1% stillborns; one female child, age - one aqueous phosphomolybdic acid followed year; three male adults of ages 27, 38, by mordanting in 5% phosphotungstic and 42 years; and three female adults of acid (Romeis, '40). ages 37, 39 and 62 years. The postmortem 3. 2% orange G in 5% aqueous phos- interval preceding fixation was between photungstic acid (Ezrin and Murray, '63). three and five hours. Two additional adult 4. 0.3% acid fuchsin (C.I. 42685) in pituitaries, fixed after ten and fourteen 1% acetic acid. hours, were utilized to evaluate fixation 5. 0.7%ponceau R (C. I. 16150) in 1% but were not employed in the cytological acetic acid. studies. 6. Masson A (a mixture of acid fuchsin When possible the glands were divided and ponceau R, as above), with and with- sagittally prior to fixation, and both frontal out post-mordanting in Masson B (1% and sagittal sections were obtained from aqueous phosphomolybdic acid) (Masson, the same gland in order to have a basis '29). for determining cellular distribution. The 7. 2% aniline blue WS (C. I. 707) in material was fixed in 10% neutral fonna- 2% acetic acid (Masson C). lin (19 pituitaries) or Bouin's fixative (5 8. 2% light green SF yellowish (C. I. pituitaries). Formalin was utilized be- 42095) in 1% acetic acid. cause an evaluation of staining procedures 9. 1% aqueous erythrosin B (C. I. after this fixative would be most useful 45430). to other investigators who have access to 10. Mallory I1 (Humason, '62) (0.5% existing, usable material which has been aniline blue and 2% orange G in 1% routinely fixed in formalin. Bouin's fixa- acetic acid). tive was employed in order to have a 11. 0.1% luxol fast blue MBSN (lot no. means of evaluating cellular preservation 136, E. I. Dupont Co.) in 95% alcohol by a fixative other than formalin. (Kluver and Barera, '58). The staining procedures which were 12. 0.5% aqueous acid alizarin blue 2B evaluated were those previously reported (C. I. 1063) (Humason, '62). by a number of investigators. These pro- cedures were employed, where possible, as These procedures were employed singly, originally reported and were also modified in combination with each other (table l), as needed. In methods which utilized a and with the following stains for mucoid cells : aldehyde fuchsin, resorcin fuchsin, sequence of stains, the stains were em- alcian blue, periodic acid Schiff (PAS), ployed singly and in step-wise order, to be aldehyde thionin and colloidal iron. The certain of the identity of the stained cells. details of the mucoid procedures and the As an illustration, in the use of the Mas- oxidation methods employed have been son trichrome method after aldehyde fuch- summarized elsewhere (Conklin, '67). sin different sections were stained in (1) aldehyde fuchsin (ald. f.); (2) Masson A Several structural or tinctorial qualities (Mas. A); (3) Mas. A, B; (4) Mas. A, B, were used to identify several cell types. These qualities were : light green (It. gr.); (5) It. gr., ald. f., (1) the staining reaction intracyto- Mas. A; (6) ald. f., Mas. A, B; (7) ald. f., of plasmic granules; Mas. A, B, It. gr. A similar procedure was (2) the staining reaction of cytoplasm utilized with all of the other methods em- when granules were absent; ployed. (3) the morphology of the cells, espe- The procedures for acidophilic cells cially with regard to nuclei and cytoplas- which were evaluated are summarized below: 'The material utilized in this study was obtained 1. 0.1% azocarmine G (C. I. 50085) in from the Department of Pathology and from the Embryology Research Collection of the Department 1% acetic acid at 60°C preceded by and of Anatomy. PITUITARY ACIDOPHILS 349

TABLE 1 Summary of methods employed

Azocarmine erythrosin, Mallory I1 Orange G erthrosin, Mallory 11, acid alizarin blue (Helant, '60; Racadot, '62) Aniline blue R'Iallory I1 (Humason, '62) Light green azocarmine, orange G (Romeis,'40) Acid fuchsin azocarmine, orange G, light green Ponceau R 1 azocarmine, orange G, aniline blue (Romeis, '40) Masson A azocarmine, Masson B, C Erythrosin B orange G, light green Lux01 fast blue Masson A, B, C (Masson, '29) Acid alizarin blue Masson A, B, light green Masson A, B Masson B, C

1 These stains were employed with and without mordanting in Masson B. mic granules with little reliance on size Mucoid cells were those whose granules and shape of the cells; were stained by mucoid procedures such (4) cellular distribution within the pars as PAS, alcian blue or aldehyde fuchsin. distalis and the nature of the distribution, There are actually two groups of cells in i.e., whether the cells occurred singly or the mucoid category, i.e., cells whose gran- in clumps. ules stained only with mucoid procedures Adjacent sections were sometimes (mucoid cells) and cells whose granules stained by different methods and cell were stained by mucoid procedures and counts were employed to aid in the identi- acidophilic dyes (acidophilic-mucoid cells). fication of cell types. Cell counts were The degranulated cells were those that made by inserting a disc micrometer ruled possessed morphological characteristics into 49 squares into the ocular of the that permitted classifying them either as microscope and counting a minimum of acidophilic or mucoid, but which were 1,000 cells in a specific area of the gland. lacking in specific cytoplasmic granules. Comparison of the cell counts assisted in It was considered that these and the modi- evaluating the specificity of certain stain- fied cells were degranulated forms of ing reactions. either acidophilic or mucoid cells. Various terms have been employed to The final chromophilic cell types were indicate different classes of pituitary cells the modified acidophilic and mucoid cells. and also to identify individual cells within These could be classified as either acido- these classes. Romeis ('40) originally di- philic or mucoid because either a few vided the cells of the pars distalis into cytoplasmic granules or vesicles were pres- three groups, the acidophils, basophils ent that exhibited specific staining char- and chromophobes. Later, Pearse ('52a) acteristics. Included in this group were employed the term mucoid to identify the the beta 3 cell of Ezrin and Murray ('63), basophils of Romeis since the cells in this the Crooke-Russell and Crooke hyaline group exhibited some degree of staining cells, the gamma cell (Romeis, '40), and after the PAS reaction. More recently, the the vesiculate mucoid cell (Pearse, '52b). term mucoid cells has included cells which In the present study, chromophobic cells are stained by all methods €or muco- and were identified by the absence of resolv- glycoproteins. able, cytoplasmic granules. Chromophobes In the present report, the chromophilic were small and the nucleus was enveloped cells of the pars distalis were classified as by a thin rim of cytoplasm (fig. 12). The either acidophilic, mucoid, degranulated nuclei were usually oval, contained a or modified. The acidophils included small nucleolus and exhibited a variable those cells which exhibited an affinity for chromatin pattern. When the cells were acid dyes and were not stained by mucoid of sufficient size to exhibit any specific stains except after certain oxidative pro- staining response they were classified as cedures (Conklin, '67). a type of chromophil. 350 JAMES L. CONXLIN RESULTS +++++ + Examination of the fol- ++++++++ lowing the application of a given acido- ++++-I- + philic stain revealed that the granules of most cells were stained to some degree. Therefore, identification of specific cell ++ + types necessitated the use of several stain- ++++++++++ ing procedures since with few exceptions ++ + specificity was unattainable with a single technique. When the staining reactions of pituitary cells are described, reference to a specific cell is based on its identification after a variety of procedures. Individual cells have been referred to by a variety of terms including alphabeti- cal designations. Recently, it has been +++++ N+ recommended that a functional terminol- ++++++++ ogy be employed for pituitary cells in the +++++ -t absence of an agreeable morphological ter- minology (van Oordt, '65). This can be done if a technique is employed which gives functional results. However, since ++ it was necessary to employ some designa- tion for the different cell types in the present report, the system of Ezrin ('63) p1 which is analogous to the terminology of ++ + Romeis ('40) was employed. According to + + + + + + + + this system the chromophilic cells are in- ++ + dicated by Roman numerals I-VIII. Of these the definitive acidophils are types I, 11, and VIII which correspond respectively ++ + t- to the alpha, eta and epsilon cells of ++++++++ Romeis. Cell types 111-VII are mucoid ++ + + cells which include the amphophils (Burt and Velardo, '54), beta, delta and gamma cells. The staining characteristics of nine cell + types are described (table 2). Cell type ++++ ++++ IX is a mucoid cell found in the posterior + zone of the pars distnlis which differs slightly from the anterior zone mucoid .. cells. A blank space in the table is an ++++ -t indication that a cell type either did not ++++++++ + exhibit an affinity for a stain or was only ++++ + weakly stained. Since most cell types were stained by one or more of the acid stains, identifica- tion of acidophils was dependent upon determining which of the cell types were also mucoid cells. Application of mucoid staining procedures revealed that cell types 111-VII and IX were stained by these methods (Conklin, '67). Of this group, cell types 111, IV, VI, and IX were clearly acidophilic-mucoid cells. Therefore, of the PITUITARY ACIDOPHILS 351 nine cell types only types I, I1 and VIII They were most numerous in the posterior were distinct acidophils and are described region of the gland, anterio-lateral to the in detail in this report. remnants of the residual cleft. Large clus- ters of the cells also occurred in the Characteristics of the acidophils central region of the gland around the Specific identification of the three types connective tissue septum. In the anterior of acidophilic cells was possible when cer- region of the gland they were more nu- tain acid stains were employed in conjunc- merous inferiorly than superiorly. Very tion with procedures for mucoid cells few other cell types were observed in the (table 3). acidophilic zones and the cells occurred in large irregular clusters in close association Cell Type I with sinusoids. The typical, or most ob- The type I cell (figs. 2-8) contained served, type I cell ranged in size from characteristically small, densely packed small round cells (fig. 5) with scanty intracytoplasmic granules. The granules cytoplasm and a central nucleus to large were specifically identifiable by being car- oval cells 20-25 c1 that contained abun- minophilic following an azocarmine-or- dant cytoplasm and an eccentric nucleus ange G sequence of staining. The gran- (figs. 2-4). The large cells were often ules were unstained by mucoid stains binucleate and most of them exhibited a although the cytoplasm of the cells was sparsely granulated, juxtanuclear zone (fig. lightly PAS positive. The cells were pres- 4) and often contained 1-2 intracytoplas- ent throughout the anterior zone and were mic vacuoles. The nuclei of the large rarely present in the posterior zone (fig. 1). type I cell exhibited some morphological

TABLE 3 The tinctoriul properties of acidophit cells

Staining Cell type procedures employed I I1 VIII PAS pale pink pale pink pale pink Performic acid,alcian blue, PAS pale pink pale pink pale pink Performic acid,alcian blue, PAS, orange G dark yellow pale yellow dark yellow Aldehyde thionin, PAS pale pink pale pink pale pink Aldehyde thionin, PAS, orange G dark yellow pale yellow dark yellow Colloidal iron, PAS, orange G dark yellow pale yellow dark yellow Herlant tetrachrome pale orange grey cytoplasm, light orange red granules cytoplasm, pale red granules Aldehyde fuchsin, without oxidation, Romeis counterstain red red-orange red-orange Aldehyde fuchsin, without oxidation, Masson trichrome red red-orange red-orange Aldehyde fuchsin, periodic acid oxidation, Masson trichrome red red-orange red-orange Performic acid, alcian blue, aldehyde fuchsin light purple light purple light purple 352 JAMES L. CONKLIN variation. Although most nuclei were ves- guishing characteristics were presented by icular with a fine chromatin pattern and the nuclei of the type I1 cells. The cells a prominent basophilic nucleolus, some were rarely binucleated; the chromatin cells contained a dark almost-pyknotic nu- was coarse and the nucleolus was promi- cleus (fig. 6). The nucleolus in the latter nent. The nuclei were either centric or type of cell was usually acidophilic. The eccentric in location and when in the lat- cytoplasmic granules of the darkly nu- ter position a small nongranulated, juxta- cleated cells were intensely carminophilic nuclear zone was evident in the cytoplasm. and the juxtanuclear zone was usually not Type I1 cells, although variable in num- evident. For purposes of reference three ber, were observed in the pituitary gland type I cells were distinguished. These of both sexes. They were most frequent in were the small type I cell (with reference the fetal and post-menopausal pituitary. to size) (fig. 5) and the light (figs. 2-4) and dark type I cells (fig. 6) (with refer- Cell Type VIII ence to nuclear staining). The type VIII cell (figs. 14-16) was Type I cells were present in all the round to oval in shape and ranged from pituitaries examined from both fetuses and slightly larger to slightly smaller than the adults. Small and light type I cells were type I cell. The nucleus was either cen- most numerous in younger subjects while tric or eccentric in position depending after forty years of age there was an in- upon the shape of the cell. The larger, crease in the dark type I cells. Concomi- oval cells occasionally exhibited a small tant with the increase in dark cells, the juxtanuclear zone. The nucleus was ves- membranes of the cells became less dis- icular with moderately coarse chromatin tinct and the cells occurred in large syn- and usually contained a small nucleolus. cytial-like clusters. Occasionally, oddly The cytoplasm of the type VIII cell was shaped type I cells were observed in the often poorly preserved (fig. 14) and some- pituitaries of older subjects (figs. 7-8). times fragmented. This occurred after fix- The significance of these cells was not ation in either formalin or Bouin's fluid. determined. A small juxtanuclear zone was sometimes observed in the larger, oval cells. The Cell Tgpe I1 granules of the type VIII cell were mor- While the granules of the type I1 cells phologically distinct because of their large were colored by most acid stains only size and sparseness. They were stained erythrosin stained them with an intensity by most acid dyes but exhibited the great- which permitted their being distinguished est affinity for orange G and ponceau R. on the basis of color (figs. 9-11, 13). The cells were observed in all pituitary However, the cells could additionally be glands. They were located individually in identified by the sparseness and coarse- all areas of the anterior lobe including the ness of the intracytoplasmic granules. The acidophil zones and usually were in close cells were scattered throughout the gland association with the sinusoids. Type VIII although they occurred most frequently cells were easily discriminated from the near the periphery, along the superior type I cells because of the marked differ- margin. They were present most fre- ence in granule size and content. quently as single cells intermingled with It was difficult to distinguish between mucoid cells. 'Usually, the type I1 cell was the type I1 and type VIII cells after routine smaller (15-20 p) than the type I cell and procedures although it could be accom- ranged from round to pyramidal in form. plished on the basis of difference in gran- Occasionally, a type I1 cell was larger than ule size. After the Herlant procedure the a type I cell but this was observed only cells were more easily discriminated by under certain conditions (fig. 13). The the greater erythrosinophilia of the gran- cytoplasm of the type I1 cell was finely ules in the type I1 cell. granular and lightly acidophilic and was colored a light grey after the Herlant pro- DISCUSSION cedure, the result of light staining with When a single stain is employed it is aniline blue and orange G. Few distin- extremely difficult to distinguish between PITUITARY ACIDOPHILS 353 the different types of acidophils. Little not. The non-fluorescent acidophils would specificity is exhibited by acidophilic undoubtedly include the type I1 and type stains and the identification of individual VIII cells observed in this study. The Her- cell types requires the employment of a lant tetrachrome procedure clearly reveals sequence of stains. While ShankLin et al. two additional acidophilic cell types in the ('59) described luxol fast blue as a spe- human pituitary and they have also been cific stain for alpha cells it had an affinity studied by electron microscopy (Pasteels, for all acidophils. Erythrosin was differ- '63 ) . entially specific for the type I1 cell but it The type I1 cell exhibited a somewhat also stained the type VIII cell to some specific affinity for erythrosin and thus degree. Azocarmine, when employed with was tinctorially distinguished from the orange G, intensely stained the type I other cells. The type VIII cell stained with cells but again some measure of staining less specificity but was morphologically was exhibited by other acidophils and the distinguishable. The type I1 and type VIII acidophilic-mucoid cells. cells correspond respectively to the eta The true acidophils included at least and epsilon cells of Romeis ('40) (table 4). three types of cells. The carminophilia, Although Romeis did associate the epsilon oval shape and fine granulation of the cell with a specific hormone these cells type I cell (figs. 2-8) corresponded closely were always present in all pituitaries. Eta to the described by Romeis (type 11) cells occurred only in the last ('40). Also, in agreement with Romeis, trimester of pregnancy and were suggested small, light and dark alpha cells were to be the source of a fetal . observed (figs. 4-6). Romeis noted little In the present study, both type I1 and correlation between the kind of type I type VIII cells were observed in all pitui- (alpha) cell and the physiological state taries. Type I1 cells occurred most fre- of the subject. In the present study the quently in pituitaries of a term fetus (fig. small and light type I cells were predomi- 13) and a post-menopausal female. Pas- nant in the young while the dark type I teels ('63) has also reported the presence cell was more frequent in older subjects. of the eta cell in a fetal pituitary. The Most investigators of hormone localiza- failure of Romeis to observe ''pregnancy" tion have not distinguished between the or eta (type 11) cells in non-pregnant alpha cell and other acidophils. In not subjects may be the result of the close one of the several immunofiuorescent similarity in tinctorial properties of the studies (Leznoff et al., '60; Meneghelli and eta and epsilon (type VIII) cells. Scapinelli, '60) has a specific correlation The function of the eta and epsilon cells been made between the alpha cell and the is uncertain. As the result of transplanta- cells which are fluorescent. However, most tion studies Herlant ('60, '63) and Pas- investigators (see reviews) conclude that teels ('61) have concluded that erythro- the type I (alpha) cell is the probable sinophilic cells are the source of prolactin source of somatotropic hormone (STH) . in rodents. Later, Pasteels ('63) demon- Meneghelli and Scapinelli ('60) observed strated the ultrastructural similarity of eta that while most acidophils were fluores- cells in the rodent and human hypophysis. cent with anti-STH a few acidophils were Although Emmart et al. ('63) reported

TABLE 4 Classification of acidophils

Author System Terminology Romeis ('40) Greek alpha eta epsilon Ezrin ('63) Roman type I type I1 type VIII van Oordt ('65) functional somatotropic cell lactotropic cell (tentative) This report Roman, type I type rI modified cell descriptive 354 JAMES L. CONKLIN the immunofluorescent localization of pro- dans le glande pituitaire. Bull. Micr. appl., lactin in the rodent they did not correlate 10: 37-44. this localization with any specific cell type. 1963 Apport de la rnicroscopie elec- tronique a l'etude du lobe anterieur de l'hy- The higher incidence of the type I1 cell pophyse. In: Cytologie de l'adenohypophyse. in the last trimester of pregnancy in both Ed. by J. Benoit and C. Da Lage. Paris. Edi- mother and fetus is suggestive of a lacto- tions of C.N.R.S., pp. 73-90. tropic function. 1964 The cells of the adenohypophysis The function of the epsilon (type VIII) and their functional significance. In: Inter- nat'l. Rev. Cytol. Ed. by G. H. Bourne and cell remains obscure. Purves ('61) and J. F. Danielli. Academic Press, New York, Halmi ('63) have suggested that it is an 17: 299-382. inactive eta cell, while Herlant ('64) pro- Humason, G. H. 1962 Animal Tissue Tech- posed that this cell elaborated the hor- niques. W. H. Freeman and Company, San mone, ACTH. Tinctorially and morpho- Francisco. logically the type VIII cell is intermediate Kluver, H., and E. Barrera 1958 A method for in properties between the type I and type the combined staining of cells and fibers in the nervous system. J. Neuropathol. Exp. I1 cells. Until further evidence is available Neurol., 12: 400-403. the type VIII cell is best classified as a Leznoff, A., J. Fishman, L. Goodfriend, E. Mc- modified form of one of the other specific Garry, J. Beck and B. Rose 1960 Localization cell types. of fluorescent antibodies to human growth hormone in human glands. ACKNOWLEDGMENT Proc. SOC.Exp. Biol. Med., 104: 232-235. Masson, P. 1929 Some histological methods: The author is indebted to Mrs. Lana Trichrome stainings and their preliminary Brock, Mrs. Constance Miller and Miss technique. J. Techn. Methods, 12: 75-90. Martha Marsh for assistance in prepara- Meneghelli, V., and R. Scapinelli 1961 L'ormone tion of the material and manuscript. somatotropo umano e prodotto dalle cellule acidofile preipofisarie. Atti Della Societa Med- LITERATURE CITED ico-Chirurgica de Padova, 36: 21-28. Pasteels, J. L. 1961 Secretion de prolactine Burt, A. S., and J. T. Velardo 1954 Cytology par l'hypophyse en culture de tissus. C. R. of the human adenohypophysis as related to Acad. Sci. (Paris), bioassays for tropic hormones. J. Clin. Endocr., 253: 2140-2142. 14: 979-996. 1963 Etude experimentale des cellules responsables de la secretion de prolactine chez Conklin, J. L. 1967 A survey of histochemical procedures for the demonstration of mucoid les niammiferes. In: Cytologie de l'adenohy- cells in the human pars distalis. (In manu- pophyse. Ed. by J. Beniot and C. Da Lage. script. ) Paris. Editions of C.N.R.S., pp. 137-148. Emmart, E. W., S. S. Spicer and R. W. Bates Pease, A. G. E. 1952a Observations on the 1963 Localization of prolactin within the localization, nature and chemical constitution pituitary by specific fluorescent antiprolactin of some components of the anterior hypophy- globulin. J. Histochem., 11: 365-373. sis. J. Path. and Bact., 64: 791-810. Ezrin, C. 1963 In: Discussion Generale, Cytob - 1952b The cytochemistry and cytology ogie de l'adenohypophyse. Ed. by J. Beniot and of the normal anterior hypophysis investigated C. Da Lage. Paris. Editions of C.N.R.S., p. 346. by the trichrome-periodic acid Schiff method. - 1964 The pituitary gland. Ciba Clini- J. Path. and Bact., 64: 811-826. cal Symposia, 16: 71-100. Purves, H. D. 1961 Morphology of the hypoph- Ezrin, C., and S. Murray 1963 The cells of the ysis related to its function. In: Sex and In- human adenohypophysis in pregnancy, ternal Secretion. Ed. by W. C. Young, The disease and adrenal cortical disorders. In: Williams and Wilkins Company, Baltimore. Cytologie de I'adenohypophyse. Ed. by J. 1: 161-239. Beniot and C. DaLage. Paris. Editions of Purves, H. D., and E. G. Bassett 1963 The C.N.R.S., pp. 183-200. staining reactions of the pars intermedia cells Halmi, N. S. 1963 Some unsolved problems of and their differentiation from pars antedor anterior pituitary histophysiology. In: Cytol- cells. In: Cytologie de l'adenohypophyse. Ed. ogie de l'adenohypophyse. Ed. by J. Benoit and by J. Benoit and C. Da Lage. Paris. Editions C. DaLage. Paris. Editions of C.N.R.S., pp. of C.N.R.S., pp. 231-242. 19-32. Racadot, J. 1962 Sur La Mise En Evidence Des Herlant, M. 1960 Etude critique de deux tech- Types Cellulaires Adenohypophysaires Par La niques nouvelles distinies a niettre en evidence Methode De Herlant Au Bleu D'Alizarine Acide les differentes categories cellulaires presentees Bull. Micr. appl., 12: 16-20. PITUITARY ACIDOPHILS 355

Romeis, B. 1940 Inner sekretorische Drusen alpha cells in the human pituitary. Stain 11. Hypophyse. In: Handbuch der Microskopi- Technol., 34: 55-58. schen Anatomie des Menschen 6. Ed. by W. van Oordt, P. G. W. J. 1965 Nomenclature of von Mollendorf, published by Julius Springer, the hormone producing cells in the adenohypo- Berlin. physis. A report of the International Com- Shanklin, W. M., T. K. Nassar and M. Issidores mittee for Nomenclature of the Adenohypophy- 1959 Lux01 fast blue as a selective stain for sis. Gen. Comp. Endocr., 5: 131-134. PLATE 1

EXPLANATION OF FIGURES

1 The pars distaEis (PD) and pars nervosa (PN) of the human pituitary gland. (AZ) anterior zone and (PZ) posterior zone of the pars distalis. The dark areas in the center are colloid filled remnants of the residual cleft. Aldehyde fuchsin and Masson. x 200. 2 A portion of an acinus containing oval and pyramidal type I cells. Note the relation to the sinusoid(s). Aldehyde fuchsin and Romeis’ azan. x 2000. 3 A variant morphological form of the type I cell. Note the numerous vesicles (light areas) in the cytoplasm. Aldehyde fuchsin and Masson. Wratten no. 22 filter. X 2600. 4 An oval type I cell which contains a prominent justanuclear area. Note the density and size of the cytoplasmic granules in this and the other figures. The nucleus is typical of the light type I cell. Aldehyde fuchsin and Romeis’ azan. Wratten no. 22 filter. X 2600. 5 A group of small type I cells. Herlant tetrachrome. X 2300. 6 A group of dark type I cells. Note the pronounced nuclear density. Herlant tetrachrome. x 2300.

356 PLATE PITUITARY ACIDOPHILS 1 James L. Conklin

357 PLATE 2

EXPLANATION OF FIGURES

7-8 Pleimorphic forms of the type I cell. Aldehyde fuchsin and Mas- son. x 2300. 9 A type I1 cell. Note the coarseness of the cytoplasmic granules as contrasted with the granules of the type I cell. Herlant tetra- chrome. Interference green filter. X 2600. 10-11 Type I1 cells after staining with Masson trichrome. X 2300. 12 Chromophobes (c) and other cell types. Note the paucity of cyto- plasm and lack of granulation in the chromophobic cells. Herlant tetrachrome. x 2000.

358 PITUITARY ACIDOPHILS PLATE 2 James L. Conklin

359 PITUITARY ACIDOPHILS PLATE 3 James L. Conklin

EXPLANATION OF FIGURES 13 A type I1 cell from the hypophysis of a term fetus. The cell contains numerous granules and is larger than the adult type I1 cell. Herlant tetrachrome. Interference green filter. X 2000. 14 A type VIII cell. The poor preservation of the cvtoplasm is characteristic of this cell. Her- Iant tetrachrome. Interference green filter. x 2000. 15-16 Type VIII cells exhibiting differences in granulation. Observe the prominent nucleolus in the cell in figure 15. Note the similarity in appearance between the cell in figure 16 and the type I1 cells. Aldehyde fuchsin and Masson. x 2000.

360