Specific ultrastructural markers of human pinealomas

Acta Neuropathologica

Acta Neuropathol (Berl) (1983) 62:31 140

9 Springer-Verlag 1983

Specific Ultrastructural Markers of Human Pinealomas A Study of Four Cases* J. H a s s o u n 1, D. G a m b a r e l l i ~, J. C. P e r a g u t 2, a n d M. T o g a 1 1 Laboratory of Neuropathology, Facult~ Timone, Bd Jean-Moulin, F-13005 Marseille, France 2 Department of Functional Neurosurgery, CHU Timone, Marseille, France

Summary. A n u l t r a s t r u c t u r a l s t u d y o f f o u r p i n e a l o m a s was c a r r i e d o u t to precise eventual specific m a r k e r s . D a r k a n d clear cells j o i n e d with z o n u l a e adherents, extensive a n d p l e i o m o r p h o u s processes, a c o m p l e x vacuolar system, and c h a r a c t e r i s t i c organelles (lysosome-like structures, clear a n d dense-core vesicles, vesicle-crowned rodlets a n d related structures, m i c r o t u b u l a r sheaves a n d c e n t r i o l a r derivatives, m e m b r a n o u s whorls, f i b r o u s bodies, m i c r o t u b u l e s , h e t e r o g e n e o u s c y t o p l a s m i c inclusions) offered a typical p a t t e r n . N o c o r r e l a t i o n c o u l d be m a d e between the histological a n d u l t r a s t r u c t u r a l features. T h e a u t h o r s stress the ultras t r u c t u r a l similarities between the h u m a n t u m o r cells a n d the m a m m a l i a n p i n e a l cells. P i n e a l o m a s a p p e a r e d as a m o r p h o l o g i c a l entity distinct f r o m n e u r o n a l a n d astrocytic tumors. K e y words: H u m a n p i n e a l o m a s Specific m a r k e r s - P i n e a l o c y t e

s o m e extent, to this o f the pineal cells o f o t h e r mammals.

Materials and Methods Tumor fragments were removed by stereotactic biopsy in four patients (Table 1). In each case, tumor fragments were fixed by immersion in 4 % glutaraldehyde, post-fixed in 1% OsO4. Following dehydratation in graded acetone, they were embedded in araldite for 1 ~tm thick and ultrathin sections. Semi-thin sections were stained with toluidine blue, hematoxylin-phloxin-saffron, Bodian method for neurofibrils and immunoperoxidase method for glial fibrillary acidic protein (GFAP) (Immunolok, Histoset). Ultrathin sections were contrasted with uranyl acetate and lead citrate, observed and photographed with a Philips EM 300 electron microscope.

Ultrastructure -

Results Light Microscopy

Introduction T h e u l t r a s t r u c t u r e o f the h u m a n a d u l t p i n e a l g l a n d is u n k n o w n . O n l y a few w o r k s have dealt with the fine structure o f the fetal p i n e a l g l a n d ( M N l e r 1974, 1976). True pinealomas consequently constitute a unique a p p r o a c h to its m o r p h o l o g y . H o w e v e r , these t u m o r s are very rare as well as the u l t r a s t r u c t u r a l w o r k s on this t o p i c (Nielsen a n d W i l s o n 1975; H e r r i c k a n d R u b i n s t e i n 1979; K l i n e et al. 1979; M a r k e s b e r r y et al. 1981). F o u r n e w cases are r e p o r t e d here. T h e u l t r a s t r u c t u r a l findings have p e r m i t t e d us to elicit a precise profile o f t u m o r p i n e a l o c y t e s w h i c h c o u l d be related, to * Supported by a grant of FEGEFLUC (Fbd6ration Nationale des Groupements des Entreprises Fran~aises darts la Lutte contre le Cancer), Marseille Offprint requests to: Dr. J. Hassoun (address see above)

The t u m o r s p r e s e n t e d similar features in all cases. N o l o b u l a r architecture was d e m o n s t r a t e d in a n y case. T h e t u m o r tissue was c o m p o s e d o f densely p a c k e d small

Table t. Patients operated for a pineal parenchymal tumor. Sex, age, surgical approach, treatment, and postoperative survey Case 1

Case 2

M, 31yr - Stereotactic biopsy, then subtotal surgical removal. Radiotherapy and chemotherapy. Residual tumor nodule on CT scan after a 19-month course F, 14 yr - Stereotactic biopsy. Radiotherapy and chemotherapy. No tumoral change on CT scan after a 21-month course

Case 3

Case 4

F, 44 yr - Stereotactic biopsy. Radiotherapy and chemotherapy, No residual tumor on CT scan after a 15-month course F, 72 yr -- Stereotactic biopsy. Treatment refused. No data about the clinical course

32

J. Hassoun et al: Ultrastructure of Pinealomas

Fig, ! a, b. Histologicalaspectsof pinealomas,a Rosette-likearrangementof tumors cells ( x 212, case2). b Extensivefibrillaryand vacuolated stroma ( x 350, case3~

round cells in a more or less abundant fibrillary stroma (Fig. 1). Nuclear chromatin was finely punctuated. Mitoses were very rare. The cytoplasm was scanty, often clear. In some places, tumor cells formed concentric arrangements evoking rosettes (Fig. la). Other areas showed a more extensive, often vacuolated stroma (Fig. 1 b) particularly in case 3. Blood Capillaries were unremarkable. Rare G F A P positive protoplasmic astrocytes were found in all cases, but tumor cells were always negative.

Electron Microscopy All the tumors were composed of a variable proportion of clear and dark cells joined with extensive zonulae adherentes and presenting pleiomorphous intermingled processes (Figs. 2, 3, 4a). Nuclei with clumped and marginated chromatin were round or indented (Fig. 2). Nucleoli were unconspicuous. In perikarya the most frequent organelles corresponded to numerous irregular and osmiophilic lysosome-like inclusions (Figs. 3b, 4b), microtubules (Fig. 4d) and dense-core vesicles (DCV) (Fig. 4a, c). Two populations of DCV were observed: 8 0 - 1 0 0 n m in all cases and 2 0 0 230 nm in cases 2 and 3. Extensive smooth reticulum

was found in all cases with peculiar arrangements as concentric whorls (Fig. 4e), piled-up cisternae and annulate lamellae. Clusters of centrioles and sparse cilia (Fig. 4b) appeared as other constant characteristics. In three cases (2, 3, and 4) microtubular sheaves (MTS) were observed close to centrioles in Golgi area (Fig. 6 a-d). On longitudinal sections they were formed by straight or slightly curved microtubules ensheathed in an osmiophilic material. Structures composed of three or four doublets of microtubules forming a hemicentriole were observed in some zones (Fig. 6 e). They were interpretated as transversal sections of MTS. Filaments were exceptionally found in otherwise typical tumor cells; at the opposite, 0 . 5 - 0 . 8 g m ovoid "fibrous bodies" composed of short slightly waved filaments and devoid of limiting membrane were observed in three cases (2, 3, and 4) (Fig. 4d). Glycogen particles were diffuse in some perikarya. Cell processes (Fig. 1 a) contained numerous microtubules, aggregates of mitochondria, clusters of clear synaptic-like vesicles (Fig. 4a), rarer DCV (Fig. 2b) and multivesicular bodies. In some places, dark cells displayed microvilli and or loose thin processes containing DCV and projecting into the intercellular space or occupying the center of a rosette-like arrangement (Fig. 2b). Cell processes

J, Hassoun et al: Ultrastructure of Pinealomas

33

Fig. 2a, b. General view ofpinealomas, a Tumor cells with round nuclei and narrow cytoplasmic rims, inside a dense neuropile ( x 11,400, case 1). b Rosette-like arrangement of minute tumor cell processes ( x 11,400, case 2)

34

J. H a s s o u n et al: Ultrastructure of Pinealomas

Fig. 3 a, b. Vacuolar system in pinealoma, a Vacuoles delineated by both dark and clear t u m o r cells or their processes ( x 6,460, case 3). b Vacuoles appear here rather as intra-cytoplasmic structures. Note n u m e r o u s lysosome-like inclusions ( x 11,400, case 3)

Fig. 4a--e. Ultrastructural details, a Dark and clear cells joined with a zonula adherens (arrow). Note clear and dense-core vesicles in processes ( • 26,000, Case 2). b Transverse section of two atypical intra-cytoplasmic cilia in a dark cell. Numerous dense mitochondria, Golgi complex and lysosome-like inclusions ( x 34,200, case3), e DCV originating in Golgi cisterns ( • 34,200, case2), d Fibrous body (star) and microtubules ( x 41,800, case 3). e Membranous whorl ( • 34,200, case 2)

36

J. H a s s o u n et al: Ultrastructure of Pinealomas

Fig. 5a--i. V C R and related structures, Case 3 : clusters of V C R a r o u n d vacuoles (a x 34,200 - arrows; c x 41,800 - arrows; tl x 34,200), in cytoplasm (b x 34,200) and in processes (e x 34,200; f x 41,800). Case 2: structures related to V C R : osmiophilic rodlets (g and h x 41,800) and incomplete "synaptic" spherules (i • 41,800)

J. H a s s o u n et al: U l t r a s t r u c t u r e o f P i n e a l o m a s

37

F i g . 6 a - - i . M T S a n d related c e n t r i o l a r derivatives (a x 41,800, c a s e 3 ; b x 34,200, c a s e 2 ; e x 34,200, c a s e 4 ; d x 41,800, c a s e 3 ; e • 41,800 arrow, case 3). H e t e r o g e n e o u s c y t o p l a s m i c i n c l u s i o n s (f • 24,000, case 4; h • 41,800, case 4; i • 4t ,800 - fibrillary c o m p o n e n t - arrows, case 3)

a n d n e u r o m e l a n i n - l i k e inclusions (g x 20,900, case 4)

38 joined with zonulae adherentes delineated small round cavities and could form invaginations inside the tumor cells themselves (Fig. 3 a). On the other hand, simple clear-cut cavities were found in the cytoplasm of some tumor cells (Fig. 3b): they were electron-lucent or contained membranous profiles and a finely granular material. It was often impossible to precise their origin from the reticulum channels or from the extracellular space. Finally, cases 2 and 3 were characterized by the high frequency of vesicle-crowned rodlets (VCR) and related structures (Fig. 5). Typical VCR were numerous in case 3, less frequent in case 2. They were composed of one or several osmiophilic straight or curved rodlets, 100-200nm in length, 50nm in width, and of 2 0 40 nm clear vesicles (Fig. 5 a, b). In very rare examples, the rodlets showed a trilaminar substructure. In both cases, related structures corresponded either to osmiophilic rodlets not associated with vesicles but with a finely granular matrix (Fig. 5g, h) or to osmiophilic illdefined 150nm spherules (Fig. 5i). VCR and related structures were observed in both perikarya and cell processes. More often they were perpendicular or parallel to the plasma membrane. In case 3, they were predominantly observed around intra- or extracellular vacuoles; this case was also very rich in clusters of VCR, located inside the cytoplasm (Fig. 5b) or the processes (Fig. 5e, f) without any obvious relationship with the plasma membrane. Heterogeneous cytoplasmic round or ovoid inclusions were highly suggestive of pigment deposits in all cases (Fig. 6f, i). They displayed a finely granular or fibrillary matrix and amorphous osmiophilic globules sometimes denser at their periphery. In case 4 only, more osmiophilic and homogeneous deposits showed a reduced granular matrix and were reminiscent of neuromelanin pigment (Fig. 6 g).

Discussion

Widely accepted classifications have opposed two types of pineal parenchymal tumors, i.e., immature pineoblastomas and more differentiated pineocytomas (Russel and Rubinstein 1963; Rubinstein 1981). In a study of 28 cases, Herrick and Rubinstein (1979) isolated several subgroups in each type according to the cytologic maturation of the tumor cells toward photoreceptor, astrocytic and/or neuronal cells. By light microscopy, the present cases were difficult to situate inside one of these precise subgroups: they all presented transitional patterns between Homer-Wright rosettes of pineoblastomas and larger fibrillary rosettes of pineocytomas. The lobular architecture described in some pineocytomas was never seen, no more than the mosaic pattern of pineoblastomas evoking the fetal

J. Hassounet al: Ultrastructureof Pinealomas pineal gland. So the less precise term of pinealoma appears more convenient here. As far as we are aware, ultrastructural studies of human pinealomas are very rare in the literature. Four works have been published until now: Nielsen and Wilson (1975) (one case of pineocytoma); Herrick and Rubinstein (1979) (case no.27: pineocytoma with astrocytic and neuronal differentiation); Kline et al. (1979) (one case of pineoblastoma); Markesberry et al. (1981) (one case of pineoblastoma and one transitional form). On the other hand, isolated microphotographs concerning ultrastructural details only have been published (Neuwelt et al. 1979; Kurumado et al. 1976; Borit and Blackwood 1979). The present study of four tumors has permitted to find out several constant ultrastructural features which can be considered as specific markers for human tumor pinealocytes. The first striking finding corresponded to the presence of dark and clear tumor cells joined with extensive zonulae adherentes and lying among pleiomorphous processes. In their first case of pineoblastoma, Markesberry et al. (1981) correlated dark cells with oligodendroglia or microglia. In our material, both dark and clear cells appeared as two variants of tumor pinealocytes. However, dark cells specifically showed microvilli and/or short tapered processes. Another distinctive pattern found only in two cases was represented by concentric arrangements of both types of cells and ofj oined processes around vacuoles. Such a complex intrication of tumor cells ruled out the possibility of an edematous distension of the smooth reticulum and of the extracellular space. It could be related to the vacuolar system described in pinealocytes of some mammals (Pevet 1979, 1981). For Welsh and Reiter (1978) calcareous concretions originate within these vacuoles. Regarding the cytoplasmic organelles in the present cases, DCV and clear synaptic-like vesicles as well as VCR and MTS deserve a special comment. DCV have been mentioned by Nielsen and Wilson (1975) and illustrated by Herrick and Rubinstein (1979) and Markesberry et al. (1981) (case2). They were herein located in cell perikarya and processes and associated with clear synaptic-like vesicles and multivesicular bodies only in processes or bulbous endings. For Herrick and Rubinstein (1979), they supported the neuronal differentiation of tumor pinealocyte. However, they have been described in the pinealocytes of practically all the mammalian species studied to date (Pevet 1979) but not mentioned in human fetal pineal gland by MNler (1974). Like DCV, the clear vesicles numerous in the present cases, appear as a normal component of the mammalian pinealocyte (Karasek 1976; Karasek and Vollrath 1982). VCR also called "synaptic ribbons" have been observed in two of our

J. Hassoun et al: Ultrastructure of Pinealomas

cases. These minute organelles were difficult to find, needing an appropriate investigation. They have been described in pinealocytes of most of mammalian species, including the primate (Pevet 1979) and reported in tumor cell processes by Herrick and Rubinstein (1979) in a pinealoma. The physiologic significance of these organelles is debatable. In mammals they are considered as phylogenetic remnants of lower vertebrate pineatocytes where they play a role as photoreceptor (Wurtman et al. 1968). It is nowadays accepted that they do not play a true synaptic role in mammals and that their number depends on environmental lighting conditions (Vollrath 1973; Karasek and Vollrath 1982), sexual hormonal factor (Karasek 1976) and adrenergic innervation (King and Dougherty 1982). The structures related to the VCR were similar to the "synaptic" spherules (Hewing 1980; Karasek and Vollrath 1982). VCR and spherules are therefore logically expectable in human normal and tumor pinealocytes, without having the significance of a neuronal differentiation. At the opposite, normal or abnormal synaptic complexes described in true neuronal tumors of the brain (Rubinstein and Herman 1972; Hassoun et al. 1982) and of the cerebellum (Shin et al. 1978; Yagishita et al. 1982) were not found in the present tumors. MTS observed in three cases, have never been described in human pineal tumors but have been reported in papova-virus induced pineocytomas of the Syrian hamster (Varakis and Zu Rhein 1976). Originally described by Wolfe (1965) in the adult rat pinealocyte, they appear as a unique feature in some species and probably derive from centrioles during the cell differentiation. In our material their presence close to these organelles, highly favors this hypothesis. For Karasek (1976) and Ruiz-Navarro et al. (1982), MTS might be transformed into synaptic ribbons. Such a spatial correlation could not been made in the present cases. Other structures observed herein have been characteristically described in mammalian pinealocytes and found in some human pinealomas: the membranous whorls and annulate lamellae (Pevet 1979) have been also reported by Kline et al. (1979); glycogen granules (Romijn et al. 1976) have been cited by Nielsen and Wilson (1975) and by Markesberry et al. (1981, case 2). In human tumors, Nielsen and Wilson (1975), Kline et al. (1979) and Markesberry et al. (1981) stressed on the presence of cilia also seen in great number in the four present cases. They have been described by MOiler (1974) in human fetal pineal gland and constitute a typical but non specific organdie. Heterogeneous cytoplasmic inclusions were particularly numerous in our cases. Identical structures were observed by Kline et al. (1979) who compared them to granular bodies of fetal human pinealocytes (M~bller 1974). Nevertheless, this author described homo-

39

geneous but not crystalline or fibrillary inclusions. Most of the inclusions described here appeared also different from the neuromelanin granules of human substantia nigra and locus coeruleus (Duffy and Tennyson 1965; Moses et al. 1966; Hirano I971). In fact neuromelanin pigment shows three components, a granular matrix, a particulate material and variable clear lipid globules. Very rare inclusions in case4 agreed somehow with this description. On the other hand, heterogeneous cytoplasmic inclusions differed from lipofuscin or ceroid pigments, in spite of the pleiomorphous substructure of lipopigments (Samorajski et al. 1974). The significance of these peculiar cytoplasmic inclusions remains to be elucidated. Filaments described as "a very prominent feature" by M~bller (1974) were quite exceptional in our material. They have been variably reported in the perikarya and processes of tumor pinealocytes by Nielsen and Wilson (I975); Herrick and Rubinstein (1979); Kline et al. (1979). They were found neither by Markesberry et al. (1981) in their two cases nor by Varakis and Zu Rhein (1976) in the hamster papova-virus induced pineocytoma. They could not constitute a specific marker of human pinealomas. At the opposite, numerous fibrous bodies were observed in three of the present cases and appeared as a characteristic feature of these tumors. To our knowledge, they were neither reported in mammalian pineal gland nor in human pinealomas. They did not bear any resemblance to astrocytic microfilaments otherwise observed in the glial processes of the tumor stroma. The very rare GFAP-positive glial cells or processes found in all our cases did not permit, no more than the ultrastructural findings, to retain the possibility of an astrocytic differentiation from the tumor pineal cells. GFA-positive cells have been reported in rat (M~bller et al. 1978) and in human pineal gland (Loewenthal et al. 1982) and are to be considered as a normal component of this endocrine tissue as they are in normal human pituitary (Velasco et al. I982). To conclude, it can be asserted that, whatever the histological aspect of the pineal parenchymal tumors (i.e., pineoblastoma or mixed pineoblastoma-pineocytoma), numerous ultrastructural features constantly evoke those of mammalian pinealocyte. The association of intermingled dark and clear cells, intra- or extracellular vacuolar spaces, pleiomorphous cell processes, and the presence of some cytoplasmic organelles (DCV, clear vesicles, VCR, MTS, microtubules, membranous whorls, fibrous bodies, heterogeneous cytoplasmic inclusions) give these tumors a very specific pattern to be distinguished from that of the neuronal and glial tumors developed in the pineal area. Acknowledgements.We are grateful to Mrs. E. Pasquale, F. Bres, and Mr. R. Gochgagarian for their technical assistance.

40

References Borit A, Blackwood W (1979) Pineocytoma with astrocytomatous differentiation. J Neuropathol Exp Neurol 38:253-258 Duffy PE, Tennyson VM (1965) Phase and electron microscopic observations of Lewy bodies and melanin granules in the substantia nigra and locus coeruleus in Parkinson's disease. J Neuropathol Exp Neurol 24:398- 414 Hassoun J, Gambarelli D, Grisoli F, Pellet W, Salamon G, Pellissier JF, Toga M (1982) Centtal neurocytoma. An electron microscopic study of two cases. Acta Neuropathol (Berl) 56:151 - 156 Herrick MK, Rubinstein LJ (1979) The cytological differentiating potential of pineal parenchymal neoplasms (true pinealomas). Brain 102:289- 320 Hewing M (1980) Synaptic ribbons in the pineal system of normal and light deprived golden hamsters. Anat Embryol 159:71- 80 Hirano A (1971) Electron microscopy in neuropathology. In: Zimmerman HM (ed) Progress in neuropathology. Heineman, London, pp 1 - 6 1 Karasek M (1976) Quantitative changes in number of "synaptic ribbons" in rat pinealocytes after orchidectomy and in organ cultures. J Neural Transm 38:149-157 Karasek M, Vollrath L (1982) "Synaptic" ribbons and spherules of the rat pineal gland: day/night changes in vitro ? Exp Brain Res 46:205- 208 King TS, Dougherty WJ (1982) Effect of denervation on "synaptic" ribbon population in the rat pineal gland. J Neurocytol 11 : 1 9 28 Kline KT, Damjanov I, Moriber-Katz S, Schmidek H (1979) Pineoblastoma: an electron-microscopic study. Cancer 44:1692-1699 Kurumado K, Mori W, Matsutani M, Sano K (1976) Virus-like particles in human pinealoma. Acta Neuropathol (Berl) 35: 273 - 276 Loewenthal A, Flament-Durand J, Karcher D, Noppe M, Brion JP (1982) Glial cells identified by anti-e-albumin (anti GFA) in human pineal gland. J Neurochem 38:863- 865 Markesberry WR, Haugh RM, Young AB (1981) Ultrastructure of pineal parenchyma neoplasms. Acta Neuropathol (Berl) 55:143- 149 M~ller M (1974) The ultrastructure of the human fetal pineal gland. I. Cell types and blood vessels. Cell Tiss Res 152:13-30 M~bller M (1976) The ultrastructure of the human fetal pineal gland. II. Innervation and cell junctions. Cell Tiss Res 169 : 7 - 21 M~bller M, Ingild A, Bock E (1978) Immunohistochemical demonstration of S-100 protein and GFA protein in interstitial cells of rat pineal gland. Brain Res 140 : 1 -- 13 Moses HL, Ganote CE, Beaver DL, Schuffman SS (1966) Light and electron microscopic studies of pigment in human and rhesus monkey substantia nigra and locus coeruleus. Anat Rec 155:167- 184 Neuwelt EA, Glasberg M , Frenkel E, Kemp Clark W (1979) Malignant pineal region tumors. A clinico-pathological study. J Neurosurg 51 : 597- 607

J. Hassoun et al: Ultrastructure of Pinealomas Nielsen SL, Wilson WB (1975) Ultrastructure of a "pineocytoma". J Neuropathol Exp Neurol 34:148-158 Pevet P (1979) Secretory processes in the mammalian pinealocytes under natural and experimental conditions. Prog Brain Res 52:149-194 Pevet P (1981) Ultrastructure of the mammalian pinealocyte. In: Reiter RJ (ed) The pineal gland, vol 1. Anatomy and biochemistry. CRC Press, Boca Raton, pp 121 - 148 R0mijn HJ, Mud MT, Wolters PS (1976) Electron microscopic evidence of glycogen storage in the dark pinealocytes of the rabbit pineal gland. J Neural Transm 38:231-237 Rubinstein LJ, Herman MM (1972) A light and electron microscopic study of a temporal lobe ganglioglioma. J Neurol Sci 16:27- 48 Rubinstein LJ (1981) Cytogenesis and differentiation of pineal neoplasms. Human Pathol 12:441-448 Ruiz-Navarro A, Blance-Rodriguez A, Gasquez-Ortiz A, JoverMoyano A (1982) Synaptic ribbons in pinealocytes of castrated rates and rats treated with estradiol. Cell Biol Int Pep 6:629633 Russel DS, Rubinstein LJ (1963) Pathology of tumours of the nervous system. E Arnold, London, pp 173-181 Samorajski T, Ordy JM, Keefe JR (1974) The fine structure of lipofuscin age pigment in the nervous system of aged mice. In: Nanda BS (ed) Aging pigment, current research: 1. MSS Information, New York, pp 141 - 166 Shin WY, Laufer H, Lee YC, Aftalion B, Hirano A, Zimmerman HM (1978) Fine structure of a cerebeUar neuroblastoma. Acta Neuropathol (Berl) 42:11- 13 Varakis JN, Zu Rhein GM (1976) Experimental pineocytoma of the Syrian hamster induced by a human papovavirus (JC). A light and electron microscopic study. Acta Neuropathol (Berl) 35: 243 - 264 Velasco ME, Roessmann V, Gambetti P (1982) The presence of glial fibrillary acidic protein in the human" pituitary gland. J Neuropathol Exp Neurol 41 : 150-163 Vollrath L (1973) Synaptic ribbons of a mammalian pineal gland. Circadian changes. Z Zellforsch 145:171 - 183 Welsh MG, Reiter RJ (1978) The pineal gland of the gerbil Merions unguiculatus. I. An ultrastructural study. Cell Tiss Res 193:323- 336 Wolfe DE (1965) The epiphyseal cell: an electron microscopic study of its intercellular relationships and intracellular morphology in the pineal body of the albino rat. Prog Brain Res 10:332-386 Wurtman RJ, Axelrold J, Kelly DE (1968) The pineal. Academic Press, New York London, pp 2 0 - 23 Yagishita S, Itoh Y, Chiba Y, Kuwana N (1982) Morphological investigations on cerebellar "neuroblastoma" group. Acta Neuropathol (Berl) 56 : 2 2 - 28

Received June 14, 1983/Accepted August 8, 1983