A LIGHT- AND ELECTRON-MICROSCOPIC STUDY ...

Electron Microscopy and Cell Biology

ISSN 0326 - 3142 PRINTED IN ARGENTINA

1992. 16 (1) 69-86

A LIGHT- AND ELECTRON-MICROSCOPIC STUDY OF THE PINEAL COMPLEX OF THE AMMOCO ARVA OF THE SOUTHERN AMPREY Geotria australis I I I I .1

VICTOR BENNO MEYER-ROCHOW* and DUNCAN STEWART"

*•

Experimental Zoology and Elearon Microscopy; University of the West Indies; Mona-Campus; Kingston (St. Andrew) 7; Jamaica. W. I. Department of Biological Sciences; University of the Waikato, Private Bag 3105; Hamilton; New Zealand

Short title: Pineal Complex in Larval Geoaia australis lamprey Keywords: Pineal organ, ultrastruaure, lamprey, photoreceptor, brain. ABSTRACT: The pineal complex of larval Geotria australis lampreys has been examined by light- and elearon-miCTOscopy. The complex consists of a pineal organ and a smaller parapineal organ, the former being situated dorsal to the latter. It is concluded that the pineal organ is a functioning photoreceptor, with neural and endocrine output. The parapineal appears to be a more basic neuroendocrine organ without photosensory capability. Day-night comparisons of the pineal complex show no obvious differences in ultrastruaiue; this could be related to the chronobiological role of the complex as a constant monitor of ambient light levels, rather than a mere visual receptor that changes its sensitivity at night. RESUMEN: El complejo pineal de larva Geotria australis lamprea ha sido examinado con miaoscopio de luz y elearonico. El complejo esta constituido por el organo pineal y un pequeiio organo parapineal. estando situado aquel dorsalmente con respeao a este. Se concluye que el organo pineal es un fotorreceptor activo. con una funcion neural y endocrina. El parapineal parece ser un organo neuroendocrlno mas basico sin capacidad fotosensorial. Las comparaciones diumas y nocturnas del complejo pineal no muestran diferencias obvias de estructura; esto puede estar relacionado con el papel cronobiologico del complejo como un control constante de los niveles de luz ambientales, mas que a la mera recepcion visual que cambia su sensibilidad en la noche.

INTRODUCTION In lamprey ammocoetes, the pineal complex consists of a larger pineal and a smaller parapineal organ, both being evaginations of the diencephalon (Eddy. 1972; Collin et al.. 1986). The parapineal is situated ventral to the pineal in the G. australis ammocoete, although it migrates to a position rostral to the pineal at metamorphosis (Eddy and Strahan. 1970). The ammocoete pineal complex is of interest given that it remains virtually unchanged during metamorphosis, unlike the rest of the animal (Cole and Youson. 1982). The complex is already

well developed in the ammocoete, whereas the vestigial lateral eyes are still buried beneath the skin (MaskeU. 1929; Young. 1981; Tamotsu and Morita, 1986). Therefore, the pineal complex, predating the lateral eyes in the lamprey ammocoete, must be functionally important. The pineal complex is believed to act as a photoneuroendocrine organ, transducing light into neural and/or hormonal messages (Oksche, 1984; Quay. 1986; Collin et al.. 1986). The pineal complex has been found to be important in lamprey ammocoetes for synchronizing pigment migrations (Young. 1935; Eddy and

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VICTOR BENNO MEYER-ROCHOW and DUNCAN STEWART

Strahan. 1968; Eddy, 1972; Joss and Potter, 1982). locomotor activity rhythms (Hardistry. 1979), and metamorphosis (Cole and Youson. 1981). Consequently, the pineal complex has been assumed to act as a biological clock or oscillator that regulates these biorhythms (Collin et al.. 1986; Quay. 1986). Ammocoetes and adults of the southern lamprey. Geotria a astralis^ have previously been examined under the light (Dendy, 1907; Eddy and Strahan, 1970) but not the electron microscopy. From ultrastructural investigations i n other species, structural plurality of photosensory cells in the lamprey pineal complex has been reported (Meiniel. 1980; Cole and Youson. 1982), but without a knowledge of the situation in G. australis, the picture has to be naturally incomplete. An ultrastructural investigation into the G. australis complex was, thus, thought to be overdue. As no previous study of any lamprey pineal complex had specifically addressed the problem of structural or ultrastructural variations in response to the daily photocycle. this. too. was considered in the present investigation.

week later. Individual samples were then placed in fresh Zamboni's fixativefora further 2 hours, washed in several changes of buffer, and postfixed in 2% osmium tetroxlde in phosphate buffer (pH 7.4) for two hours. Subsequently, the tissues were dehydrated in a graded series of ethanol to propylene oxide, infiltrated under vacuum, and embedded in Quetol 812. Longitudinal and transverse thick sections (1 [xm), cut with glass knives, were stained with 1% toluidine blue for a few seconds on a hot plate. Photomicrographs were made using a Reichert-Jung Polyvar light microscope. Thin "silver-sections" were cut with glass and/or diamond knives on a Reichert 0M-U3 ultramiaotome and mounted on carbon-coated copper grids. Prior to examination under a Philips 400 transmission electron microscope, operated at an accelerating voltage of 80 kV, the sections were stained with saturated uranyl acetate for 20 minutes and Reynold's lead citrate (pH 12) for 7 minutes.

RESULTS Light Microscopy

MATERIAL AND METHODS Geotria australis ammocoete larvae were collected during the day from alluvial muds of the Kaniwhaniwha stream in the Waikato region of New Zealand. At the University of the Waikato the specimens were kept under a 12 h light: dark cycle, in water and substrate from the collection site at approximately IS'C. Twenty ammocoete larvae were killed by decapitation: 10 between 11.00-13.00 h and 10 between 23.00-01.00 h. Whole heads were fixed in Zamboni's modified fixative for electron microscopy, i.e. 2.1 g picric acid, 400 ml of 10% paraformaldehyde, 500 ml of 0.2 M phosphate buffer at pH 7.4, 2 ml of 25% glutaraldehyde. and 98 ml of distilled water. Finer dissection of the specimens and a further 24 hours of fixation in Zamboni's fixative (see above) followed a

Pineal organ. The gross structural arrangement of the larval G. australis pineal organ was similar to that identified by Eddy and Strahan (1970). However, the integument above the pineal (Fig. la) did not possess the slight depression noticed by them. The characteristic white appearance of the pineal causes it to be visible from the outside of the intact ammocoete. The pellucida had a dorsal curvature, but no photoreceptors. Outer segments projected into the pineal lumen. The supporting cells formed a distinct border at the edge of the lumen to give the appearance of an external limiting membrane. The ganglion cell layer connected to a pineal tract. The pineal atrium and parapineal body bore a resemblance to each other. Both had a dense layer of cells bordering the lumen, surrounded by a less

PINEAL COMPLEX IN LARVAL GEOTRIA AUSTRAUS

LAMPREY

dense layer of cells (Fig. la). There were no apparent retinomotor changes associated with the photocycle. The supporting cells of the cytoplasm ranged in length from 15-25 |im in both day and night fixed specimens. Parapineal organ. The structural arrangement of the parapineal organ was found to agree with the earlier description of Eddy and Strahan (1970), irrespective of the time of sampling. The parapineal exhibited a lumen which, depending upon the plane of sectioning, contained no outer segments, nor were there supporting cells in the epithelium. There were two epithelial layers; an inner dense layer, and an outer less dense layer with processes that projected to the lumen. The parapineal lumen was larger at the rostral end (Fig. lb). Electron Microscopy Pineal Organ Pellucida cells. The pellucida cell cytoplasm contained abundant glycogen, some Golgi bodies, smooth endoplasmic reticulum (SER), rough endoplasmic reticulum (RER), clear vesicles as well as some dense bodies. Many of the clear vesicles were found in close association with the Golgi apparatus. Some dense bodies were close in proximity to the photoreceptor outer segments and had a lamellar structure which resembled the shed outer segment debris in the lumen. Pellucida cells had either an oval or irregularly shaped nucleus. The pellucida cells frequently formed junctional complexes with the outer membranes of other pellucida cells. Photoreceptor cells. The photoreceptor cells could be divided into two main types. Type I photoreceptors were characterized by photosensitive outer segments which extended into the lumen and consisted of 70-160 lamellae each 15-20 |im long. Outer segments

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arose from a 9x2+0 cilium, which projected from the ellipsoid region of the inner segment (Figs. 2a, b). Two subgroups of type I photoreceptors were encountered. In one (Fig. 2a) the ellipsoid was packed with elongated, electron-empty modified mitochondria; in the other (Fig. 2b) less, but more typical mitochondria were present. In the first subgroup (Fig. 2a) the ellipsoid more or less abruptly turned into the myoid, which contained the typical tiny particles usually interpreted as glycogen, whereas in the second subgroup ellipsoid and myoid were connected via a narrow portion termed "isthmus" (Fig. 2c). The isthmus featured junctional complexes with the lateral plasma membranes of other photoreceptors and supporting cells (Figs. 2c, d). The myoid region contained the nucleus and some mitochondria, but particularly Golgi bodies, RER, SER, and a variety of vesicles (Fig. 2e). The basal process additionally possessed longitudinally orientated microtubules. There was some cellular variation in the presence of the different organelles in the myoid region and basal process, but the second subgroup of photoreceptor I type was in the majority. The branching synaptic processes formed thickened areas in the region of the synapse, and some interdigitated with the basal lamina. Some synaptic processes contained concentrations of clear vesicles, but obvious synaptic ribbons were not identified in these processes. The type II photoreceptors were similar to the type I, but had fewer mitochondria in the ellipsoid and myoid regions (Fig. 3a). Dense bodies similar to those in the pellucida cells were also present. The basal processes were difficult to trace, but appeared not to branch to the same degree as in the type I photoreceptors. Type II photoreceptors in the atrium had much smaller outer segments with 1-2 lamellae, 1-5 jim long, if present at all. Atrium photoreceptors also lacked an ellipsoid region (Fig. 3b, c). In rare instances, crystal conglomerates (see below) were identified in type II photoreceptors.

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Supporting cells. The supporting cells projected a 9x2+2 type cilium and numerous miaovilli into the lumen (Fig. 4a). The cytoplasm featured many large bodies, 0.6-2.0 |im in diameter, and rare rhombic crystals, 3.5-9.5 lim^ in area (Figs. 4b, c). These were sunounded by extensive tubulo-vesicular SER (Fig. 4b). Some cells also possessed an area close to the nuclei which contained ribosomes, glycogen granules, mitochondria, Golgi bodies, short SER and RER, clear vesicles as well as occasional dense bodies, and in rare instances what appeared to be crystalline conglomerates (Figs. 4d, e). Supporting cell basal processes appeared to be narrow, without branching synaptic processes. Junctional complexes of the outer plasma membrane existed between supporting cells and w i t h photoreceptors. Atrium supporting cells were smaller and clustered at the atrium edge, where junctional complexes existed with the plasma membranes of photoreceptor and other supporting cells. The nuclei of atrium supporting cells contained more heterochromatin and so appeared denser than those of the pineal retina. No dense bodies or crystal conglomerates were observed in the atrium supporting cells. Ganglion cells. The ganglion cells at the base of the pineal organ contained numerous ribosomes, presumably glycogen granules, mitochondria, Golgi bodies, RER, sparse SER, and relatively high amounts of clear and dense vesicles (Fig. 4f). The cytoplasm featured short microtubules, which became longer in the ganglion cell processes, and a concentration of clear vesicles in the axon terminals (Fig. 4f). No synaptic ribbons were identified in these ganglion cell processes. Pineal tract. The pineal tract was composed of a layer of 2-3 cells. The cells had a 9x2+2 cilium and numerous microvilli projecting from the dorsal surface into the brain cavity. Junctional complexes existed between cells at the dorsal surface of the tract. The cells

contained numerous ribosomes and presumable glycogen granules, but sparse Golgi bodies, SER, RER, mitochondria, and vesicles of any kind. Parapineal Organ Epithelial cells. There were two epithelial cell types which formed an inner electrondense and an outer electron-lucent layer. Electron-dense cells projected a single cilium and numerous microvilli into the lumen (Fig. 5a). Electron-dense cells also formed junctional complexes with the lateral membranes of electron-lucent and other electron-dense cells at the lumen edge (Fig. 5a). The cytoplasm contained mitochondria and occasionally vacuoles, but the greater electron-density stemmed mainly from a greater number of ribosomes and what seemed to be glycogen granules in the cytoplasm, and heterochromatin in the nucleus, than in the electron-lucent cells (Figs. 5a, c). The nuclei and cytoplasm were smaller, and the nuclei of a more irregular shape than in the electron-lucent cells. The basal processes were narrow, without branching processes. The outer electron-lucent layer of cells sent processes past the electron-dense cells to the lumen. These cells projected a cilium and a few microvilli, sometimes from an ellipsoid region that contained sparse mitochondria (Figs. 5b, c). Beneath the ellipsoid region was an isthmuslike region containing longitudinally-arranged microtubules, but continuous with the myoid region (Fig. 5c). This "isthmus" formed junctional complexes with other cells at the lumen surface (Fig. 5c). The myoid contained ribosomes, presumably glycogen granules, mitochondria, sparse SER fragments, clear and dense vesicles (Fig. 5d). The oval nuclei wer slightly larger than in the electron-dense cells, being 4-7 and 3-6 |am in diameter, respectively (Fig. 5d). Ganglion cells. The ganglion cells contained


LAMPREY

ribosomes, glycogen granules, clear and dense vesicles, some mitochondria, and microtubules extending into the axonal processes. The nuclei were 2.3-2.8 ^im in diameter and, therefore, considerably more voluminous than those of the epithelial cells. There were more ganglion cells in the ventral than in the dorsal parapineal region. Parapineal tract. The single layer of irregularly-shaped, almost squamous cells, projected a single 9x2+2 type cilium and some microvilli into the brain cavity. The cytoplasm contains ribosomes and possibly glycogen granules, but few other organelles or inclusions. A few cells had vacuoles. Day/Night comparison. There was no obvious difference between the day- and nightfixed specimens. There were small numbers of outer segments with expanded regions in both day and night material. There was no significant change in the diameter of the nuclei, clear and dense vesicles. Pineal photoreceptor dense bodies, except for the basal process where there was no change, were fractionally smaller at night, reducing from 0.2-0.9 to 0.2-0.4 ^im in diameter.

DISCUSSION In contrast to L. planeri (Vigh-Teichmann et al., 1989) and P. marinus (Pu and Dowling, 1981), the pineal pellucida of G. australis larvae did not possess photoreceptors. The organelles, particularly SER and clear vesicles, could by their proximity to the lumen be expected to be involved in the degradation of the photoreceptor outer segments. The dense bodies observed may be the shed lamellar structures, which have been incorporated into the pellucida. The lack of photoreceptors as well as blood vessels supports the earlier conclusion of the greater "lens-like" appearance of the G. australis pellucida (Eddy and Strahan, 1970). As a result,

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slightly more light may reach the pineal retina of G. australis than in other lampreys. Indeed, the cone-Uke pineal photoreceptor outer segments appeared to be more numerous in G. australis than previously reported in lampreys. These large outer segments may compensate for the lack of pellucida photoreceptors, so that G. australis pineal photic activity is overall similar to that of other lamprey species. Mordada mordax is thought to lack pellucida photoreceptors as well (Eddy and Strahan, 1970), but its pineal organ ultrastructure is as yet undescribed. The organelles present in the two types of pineal photoreceptors could permit active synthesis. The abundance of mitochondria in type I photoreceptors may indicate a greater energetic demand, but the branching synaptic processes are the principal evidence for suggesting that these cells are involved in neurotransmission. Type II photoreceptors form synapses, but may have an endocrine role (Eddy, 1969), given the existence of dense bodies. Serotonin has been located by fluorescence microscopy in the dense bodies of a second photoreceptor type in the pineal of L. planeri (Meiniel, 1980) with the implication that these cells are involved in indolamine metabolism (Meiniel and Hartwig, 1980). A second pineal photoreceptor type has been identified containing dense bodies in P. marinus (Cole ans Youson, 1982), and to be serotonin immunoreactive in L. japonica (Tamotsu et al., 1990). Thus, there appears to be some structural and functional specialization of photoreceptors in the lamprey pineal organ, which has been distinguished as type I photoneurosensory and, type II photoneuroendocrine cells (Cole and Youson, 1982). In the pineal organ of L. planeri, however, three types of photoreceptor cells have been identified by immuno-electron microscopy (Vigh-Teichmann et a l , 1989) and in this paper we could differentiate between two type I receptors, so that it may still be too early for a definitive classification of lamprey photoreceptors.

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photoreceptors around the atrium tend to have small or no outer segments, indicating that there is little photic activity, the size of the outer segments could be related to reduced light penetration to the region of the atrium. There may be some photic activity without outer segments in the atrium, as light sensitive cells have been identified in the brain (Hartwig and Van Veen, 1979). Similarly, the cells of the pineal tract, which in L. japonica have been likened to mammalian pinealocytes (Tamotsu and Morita, 1986) could be photosensitive; a situation also met in the Antarctic fish Pagothenia borchgrevinki (Meyer-Rochow et al.. 1988). The electron-lucent bodies in the supporting cells are thought to represent crystals which were lost during sectioning or sublimated during subsequent processing. The intact rhombic crystals are thought to be composed of guanine (Pu and Dowling, 1981) and the abundant SER surrounding the crystals has been implicated in their synthesis (Cole and Youson, 1982). The crystals reflect light (Dendy, 1907; Eddy and Strahan, 1970) and so may act as a tapetum as in the vertebrate eye (Walls, 1967). In the pineal organ a tapetum may enhance the recognition of day and night, and so help to ensure the stimulation of associated biorhythms (Quay, 1986). The organelles present in supporting cells indicate some synthetic activity, which may affect the photoreceptors through the junctional complexes. The crystal conglomerates observed could be the source of some of the dense bodies observed. Clear vesicles are far more common than the dense vesicles, especially in the synaptic processes of photoreceptors and ganglion cells. It could be that the clear vesicles have a more general role in the transport of compounds, such as neurotransmitters, and dense vesicles have a more specific, though unclarified, role (Collin etal., 1986). The parapineal organ, bearing some resemblance to the cells sunounding the atrium.

has no obvious photosensory activity. The organelles within the electron-lucent cells could indicate some secretory ability, but this is likely to be less than that of the parapineals of L. planeri and P. marinus, which have photoreceptors, some even with dense bodies (Cole and Youson, 1982; Meiniel and Hartwig, 1980). The difference is unlikely to be due to less light penetration, as the position of the parapineal organ relative to the pineal is the same in larval and adult P. marinus and G. australis (Cole and Youson, 1982; Eddy and Strahan, 1970). The extreme regression of the G. australis parapineal organ may reflect the functional dominance of the pineal pho-toreceptors which have large outer segments (see above). Recently, it has been found that serotonin induces degradation, and melatonin influences the formation, of lamellated outer segments in the pineal organ of Rana esculenta (Hartwig et al., 1991). This seems to imply that the rhythm of indolamine metabolism may be related to the cyclic renewal of the pineal outer segments. If, however, outer segment renewal in G. australis were to be a continuous process, clrcadian ultrastructural changes would be absent or, at least, less obvious. The lack of apparent ultrastructural differences in day- and nightfixed material may further be mitigated by the variation in the presence of organelles between cells of the same type. We cannot rule out that sensory cells in the pineal possess individual circadian properties, the mutual phase relationships of which could determine the nature of endogenous biorhythmlc output. In conclusion, the pineal complex of G. australis larvae appears to possess cells with photoneurosensory, photoneuroendocrine, and neuroendocrine function similar to those found in other species of lamprey. The slightly larger pineal photoreceptor outer segments may be related to the lack of pellucida photoreceptors in G. australis. The pineal supporting cells may have an as yet insufficiently understood regulatory function. In comparison to the pineal, the parapineal organ cells have the most poorly


LAMPREY

related to the lack of pellucida photoreceptors in G. australis. The pineal supporting ceUs may have an as yet insufficiently understood regulatory function. In comparison to the pineal, the parapineal organ cells have the most poorly developed ultrastructure of any lamprey yet described, but may have a small neuroendocrine capability. The apparent lack of variation in day/night conditions, particularly with regard to supporting cell crystals, may be a reflection of the pineal organ's principal role as a biological clock, rather than an additional eye.

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ACKNOWLEDGEMENTS The authors gratefully acknowledge the support and encouragement of Prof. Dr. Y. Morita and his colleagues, especially Dr. Tamotsu, of Hamamatsu Medical University, Hamamatsu, Japan, and are grateful to the staff of the New Zealand Meat Industry Research Institute, Hamilton, New Zealand, for expert; maintenance of the electron microscope used and technical advice given. Dr. J. Boubee of Electricorp New Zealand kindly helped with the fishing for specimens. Two anonymous referees provided hepful suggestions.

REFERENCES COLE WC, YOUSON JH (1981). The effect of pinealectomy, continuous light, and continuous darkness on metamorphosis of the anadromous sea lampreys Peaomyzon marinus L. J Exp Zool 218: 397-404. COLE WC, YOUSON JH (1982). Morphology of the pineal complex of the anadromous sea lamprey, Petiomyzon marinus L. Am J Anat 165: 131-163. COLLIN J-P, BRISSON P, FALCON J, VOISIN P (1986). Multiple cell types in the pineal: Functional aspects. In: Pineal and Retinal Relationships. P.J. O'Brien and D.C. Klein, Eds. Academic Press, Orlando, pp. 15-32. DENDY A (1907). On the parietal sense organs and associated structures in the New Zealand lamprey (Geotria australis). Quart J Microsc Sci 51: 1-31. EDDY JMP, STRAHAN R (1968). The role of the pineal complex in the pigmentary effeaor system of the lamprey, Mordacia mordax (Richardson) and Geotna australis fGrey). Gen Comp Endocrinol 11: 528-534. EDDY JMP (1969). Metamorphosis and the pineal complex in the brook lamprey. J Endocrinol 44: 451-452. EDDY JMP, STRAHAN R (1970). The stmcture of the epiphyseal complex of Mordacia mordax and Geotria australis (Petromyzonidae). Acta Zool 51: 67-84. EDDY JMP (1972). The pineal complex. In: The Biology of Lampreys. M.W. Hardistry and I.C. Potter, Eds. Academic Press, London, pp. 91-103. HARDISTRY MW (1979). Biology of the Cyclostomes. Chapman and Hall, London. HARTWIG HG, VAN V E E N T (1979). Spectral charaaeristics of visible radiation penetrating into the brain and stimulating extraietinal photoreceptors. J Comp Physiol A 130: 277-282. HARTWIG HG, RASCHER K, SERVOS G (1991). Autocrine control of pineal outer segment morphology in frogs. Gen Comp Endocrinol 74: 279. JOSS JMP, POTTER IC (1982). Circadian rhythms. In: The Biology of Lampreys. M.W. Hardistry and I.C. Potter. Eds. Academic Press, London, pp. 117-135. MASKELL F G (1929). On the New Zealand lamprey, Geotna australis Gray. Roy Soc NZ J 60: 167-201. MEINIEL A, HARTWIG HG (1980). Indolamines in the pineal complex of Lampetra planeri (Petromyzonidae): A fluorescence miaoscopic and microspearofluorimetric study. J Neural Transmiss 48: 65-83. MEYER-ROCHOW VB, PEHLEMANN FW (1990). Retinal organisation in the native New Zealand frogs L. archeyi, L. hamiltoni. and L. hochstetteri (Amphibia: Anura: Leiopelmatidae). J Roy Soc NZ 20: 349-366. MEYER-ROCHOW VB, VIGH-TEICHMANN I, VIGH B (1988). Elearon microscope observations on the stmaure of the pineal organ in the notothenioid fish Pagothenia borchgrevinki. In: Funaional Morphology of Neuroendocrine Systems. H.-W. Korf, B. Schaner and H.-G. Hartwig, Eds. Springer Verlag, Berlin, p. 159. OKSCHE A (1984). Evolution of the pineal complex: Correlation of stmcture and funaion. Ophthalm Res 16: 88-95. PU GA, DOWLING JE (1981). Anatomical and physiological charaaeristics of pineal photoreceptor cells in the larval lamprey, Petromyzon marinus. J Neurophysiol 46: 1018-1038. QUAY WB (1986). Pineal and biorhythms. Pineal Res Rev 4: 183-197. TAMOTSU S, MORITA Y (1986). Photoreception in pineal organs of larval and adult lampreys, Lampetra japonica. J Comp


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Physiol 159: 1-5. TAMOTSU S. KORF HW. MORITA Y, OKSCHE A (1990). Inununocytochemical localisation of serotonin and photoreceptorspecific proteins (rod-opsin, S-antigen) in the pineal complex of the river lamprey Lampetra japonica, w i * special reference to photoneuroendocrine cells. Cell Tiss Res 262: 205-216. WALLS GL (1967). The Vertebrate Eye. Hafner Publ., New York. VIGH-TEICHMANN I. VIGH B, WIRTZ GH (1989). Immunoelectron microscopy of rhodopsin and vitamin A in the pineal organ and lateral eye of the lamprey. Exp Biol 48: 203-213. YOUNG JZ (1935). The photoreceptors of lampreys 11. The functions of the pineal complex. J Exp Biol 7: 254-270. YOUNG JZ (1981). The life of vertebrates. Clarendon Press, Oxford.

FIGURAS ABBREVIATIONS A BL c cr cv db DR E ED EL G g icr I It jc

atrium basal lamina cilia crystal clear vesicle dense body dorsal retina Ellipsoid electron-dense cell electron-lucent cell ganglion cell Golgi apparatus intact crystal isthmus integument junaional complex

Lu M m mt mv Nu Os Pe PO PpO PR RER r S SER SL vSER V

VR

lumen myoid mitochondrion microtubule microvilli nucleus outer segment(s) peUuclda pineal organ parapineal organ pineal retina rough endoplasmic reticulum ribosomes supporting cell(s) smooth endoplasmic reticulum supporting-like cell vesicular SER vacuole ventral retina

Fig. 1. Longitudinal sections through the head of an ammocoete (see list of abbreviations), (a) Inset: Spatial relationship of pineal complex, brain, and integument. Bar = 100 pm. 5Qx. (b) Pellucida, lumen, and retinal epithelium of pineal organ as well as retinae of parapineal organ. Bar = 55 |jm. "SOGx.

PINEAL

COMPLEX

IN

LARVAL

GEOTRIA

AUSTRALIS

LAMPREY

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Fig. 2. Pineal organ type I photoreceptors, (a) Ellipsoid and myoid with adjacent lamellae in first subgroup, resembling conesponding region in leiopelmatid (primitivefrog)rod (see Figs. 17.18: MeyerRochow and Pehlemann. 1990). Bar - 1 pm. lO.OOOx. (b) Outer segment lamellae and ellipsoid in 2nd subgroup. Bar - 1 pm. 12.00Gx. (c) Irmer segment ellipsoid with mitochondria and Isthmus region (featuring mlaotubules) projecting through supporting cells. Bar - 1 pm. 12.000x. (d) Adjacent isthmus regions of two cells, linked by junctional complexes. Bar - 1 pm. 20,000x. (e) Iruier segment myoid containing RER and nucleus. Bar - 1 pm. lO.OOOx.

Fig. 3. Pineal organ type n photoreceptors, (a) Outer segment lamellae, a 9x2+0 type cilium (white arrow), irmer segment ellipsoids with few mitochondria and dense bodies (large anrowheads), isthmus, myoid and nuclei of pineal retina. Bar - 1 pm. 5,000x. (b) Isthmus, showing numerous miaotubules and clear vesicles Bar - 1 pm 25,000x and (c) myoid, containing SER, vesicles, dense bodies and the nucleus in the atrium. Bar - 1 pm lO.OOOx.

Fig. 4. Pineal organ supporting and ganglion cells, (a) 9x2+2 type cilium. Bar - 0.1 pm. 120,000x. (b) Supporting cell cytoplasm containing SER, spherical remnants and intaa rhombic crystals. Bar - 1 pm. 22,000x. (c) Rhombic crystal cluster. Bar - 1 pm. 3,000x. (d) Supporting cell featuring what is believed to represent Golgi and dense bodies (anowhead), Bar - 1 pm 7,500x and (e) view of perinuclear region with dense bodies and other organelles Bar - 2 pm lO.OOOx. (f) Ganglion cell wixh microtubules in the cytoplasmic and synaptic processes. Bar - 1 pm. 15,000x.

Fig. 5. Parapineal organ epithelial cells, (a) Electron dense cells with SER and clear vesicles. Note the 9x2+0 cilium (anowhead). Bar = 1 pm. 22.000x. (b) Ellipsoid region projecting a cilium and miaovilli, and containing mitochondria, SER and clear vesicles. Bar - 1 pm. 33.000x. (c) Electron-lucent myoid region with miaotubules at apical tip, RER, Golgi bodies and vesicles, projecting past elecuon-dense cell with sparse cytoplasm. Bar = 1 pm. 19,000x, (d) Myoid containing Miaotubules, vesicles, and nucleus of elearon-lucent cell. Bar = 1 pm. 15,000x.

r

M

/ .

5d

BL