Pineal Complex Research Articles

Third ventricular pineal gland grafts in the gerbil: an electron microscopical and immunohistochemical investigation.

The superficial pineal gland was grafted into the third ventricle of adult Mongolian gerbils. Donor pineal glands from both neonatal and 3-4 week old gerbils were able to survive for at least 6 months. The pinealocytes of the grafted superficial pineal glands maintained the morphology and the S-antigen immunoreactivity of the in situ pineal complex. Synaptic ribbons and spherules were present but rare. Unlike the in situ pinealocytes, glycogen accumulations were common in the graft pinealocytes. Site specific modulation of structure was indicated as the ventricular surface of the grafts became covered with cerebrospinal fluid (CSF)-contacting pinealocytes typical of those seen in the deep pineal. The CSF-contacting pinealocytes of the graft had numerous processes that extended along the ventricular surface of the graft. The blood vessels of the grafts had non-fenestrated endothelium and wide perivascular areas typical of those seen in the in situ pineal complex. Tyrosine hydroxylase-immunopositive nerve fibers were present in the grafted tissue indicating reinnervation of the graft. The source of the fibers was not determined. The nerve fibers were present both within the perivascular area and within the parenchyma where they were associated with pinealocytes. The results demonstrate that the cerebral ventricles are an ideal location for the survival of superficial pineal gland grafts. It is suggested that pineal grafts may be a means to further study pineal development and innervation.

Chapter 38: Receptor and effector mechanisms in the pineal organ

This chapter discusses a synopsis of some recent results on receptor and effector mechanisms of the pineal organ based on findings from classical studies. For several reasons, the vertebrate pineal complex—a classical circumventricular organ—appears as a valuable model to address these issues. (1) The pineal complex has undergone a conspicuous transformation in the course of phylogeny and, in comparative terms, contains sensory, neuronal, and endocrine elements. (2) The pineal comprises two very well investigated receptor-mediated signal transduction cascades—that is, photoreception and adrenoreception. (3) The pineal exerts its influences via multiple effector mechanisms—for example, propagation of electric signals via neuronal pathways as well as production and release of the neurohormone melatonin. The presence of photoreception and phototransduction in the pineal complex is clearly a plesiomorphic trait among vertebrates. The loss of a functional phototransduction cascade seems to be a mammalian apomorphy. The cladogram presents that more data on both the reptilian and the anamniotic pineal organ are needed to decide on several diverging character states, especially among birds and mammals.

Two components of the pineal organ in the mink (Mustela vison): their structural similarity to submammalian pineal complexes and calcification.

The pineal complex in the mink (Mustela vison) consists of a larger ventral and a smaller dorsal pineal. Both organs contain pinealocytes, neurons, glial cells, nerve fibers and synapses in an organization characteristic of nervous tissue. The cellular elements are arranged circularly around strait lumina. These lumina correspond to the photoreceptor spaces of submammalian pineals. A 9 + 0-type cilium marks the receptory pole of the pinealocytes which may form an inner-segment-like dendrite terminal in the pineal lumina. The cilia correspond to outer segments which form photoreceptor membrane multiplications in the pineal of submammalians and in certain insectivorous and mustelid mammals (bat, hedgehog, ferret). Axonal processes of the pinealocytes contain synaptic ribbons and terminate on intrapineal neurons of both organs. This pattern represents a neural efferentation of the pineal nervous tissue. The axonal processes of pinealocytes also form neurohormonal endings which pierce the perivascular limiting glial membrane in the ventral as well as in the dorsal pineal. The upper pineal ("epipineal") of the mink may correspond to the parapineal, frontal, or parietal organs of submammalian pineal complexes. Both pineals are encapsulated by the meningeal tissue of the brain stem. Afferent vasomotor axons of the meninges innervate smooth muscle cells of pineal arterioles. There are corpora arenacea in the pineal arachnoid and in the pineal nervous tissue, primarily in the ventral pineal. The localization of calcium ions detected around the membrane of pineal cells by pyroantimonate cytochemistry suggests membrane activity as the source of the calcium ions. The accumulation of calcium by the pinealocytes may be due to their neurosensory character. The mink is the first animal described to have both intrapineal and meningeal concrements like the human pineal.

Tyrosine hydroxylase- and neuropeptide Y-immunoreactive nerve fibers in the pineal complex of untreated rats and rats following removal of the superior cervical ganglia

The distribution of tyrosine hydroxylase (TH)- and neuropeptide Y (NPY)-immunoreactive(IR) nerve fibers in the pineal complex was investigated in untreated rats and rats following bilateral removal of the superior cervical ganglia. In normal animals, a large number of TH- and NPY-IR nerve fibers were present in the pineal capsule, the perivascular spaces, and intraparenchymally between the pinealocytes throughout the superficial pineal and deep pineal gland. A small number of TH-IR and NPY-IR nerve fibers were found in the posterior and habenular commissures, a few fibers penetrating from the commissures into the deep pineal gland. To elucidate the origin of these fibers, the superior cervical ganglion was removed bilaterally in 10 animals, and the pineal complex was examined immunohistochemically. Two weeks after the ganglionectomy, the TH-IR and NPY-IR nerve fibers in the superficial pineal gland had almost completely disappeared. On the other hand, in the deep pineal and the pineal stalk, the TH-IR and NPY-IR fibers were still present after ganglionectomy. These data show that the deep pineal gland and the pineal stalk possess an extrasympathetic innervation by TH-IR and NPY-IR fibers. It is suggested that the extrasympathetic TH-IR and NPY-IR nerve fibers innervating the deep pineal and the pineal stalk originate from the brain.

Immunohistochemical localization of somatostatin within the pineal gland of metamorphosing lampreys, Petromyzon marinus L.

The distribution and onset of immunoreactivity to anti-somatostatin-14 and anti-somatostatin-34 within the pineal gland of metamorphosing lampreys, Petromyzon marinus, were examined by immunohistochemistry. No staining was observed with anti-somatostatin-34. Somatostatin-14 immunoreactive cells were not found in the larval pineal complex but were present within the pineal pellucida of the pineal gland beginning at stage 5 of transformation and through the final two stages (6 and 7) of metamorphosis. No immunoreactivity was seen in the parapineal gland. The time of onset of this immunoreactivity in the pineal gland may denote an important transition period from larval to adult life.

Ultrastructure and opsin immunocytochemistry of the pineal complex of the larval Arctic charr Salvelinus alpinus: A comparison with the retina

The fine structure and opsin immunocytochemistry of the pineal and parapineal organs of the salmonid fish Salvelinus alpinus, the landlocked Arctic charr, were studied and compared with the retina in various developmental stages, from prehatching to two-month-old. For opsin immunocytochemistry two polyclonal antibovine rhodopsin and the monoclonal antichicken opsin antibodies OS-2 (detecting blue and green pigments) and OS-1 (detecting green and red pigments) were used. Histologically, the pineal organ consists of nervous tissue like that of the retina. It is composed of photoreceptor pinealocytes, which formed axon terminals containing synaptic ribbons, on the dendrites and perikarya of secondary pineal neurons. Already in prehatching embryos, both the pineal and retinal photoreceptors display well-developed outer segments and form synaptic terminals. The distal part of the pineal organ differentiates earlier than its proximal stalk. The differentiation of the retina starts centrally, but the caudal and dorsal retinae are differentiated earlier than the rostral and ventral ones. At the end of the larval period, the lateral retina is still undifferentiated. In all stages studied, (rhod)opsin immunoreactivity was found in the outer segments of the pineal organ and rod-type retinal photoreceptors, a finding speaking in favour of the presence of the opsin of a rhodopsin/porphyropsin. Cone-type retinal photoreceptors identified morphologically in the pre- and posthatching stages were opsin-immunonegative with the four primary antisera used. This result suggests that in the charr the opsins of cone visual pigments differ in their chemical nature from those of rhodopsin/porphyropsin. The parapineal organ was opsin immunonegative. Using the monoclonal antibody OS-2 opsin immunoreactivity was also detected in inner segments, perikarya, and pedicles of rod-type photoreceptors of both retina and pineal organ of embryos and 1- to 4-day-old larvae. This may indicate a high level of opsin gene expression during photoreceptor growth around hatching. The well-developed pineal organ and its opsin content are discussed in connection with the photonegative behaviour of the larval charr.

Efferent projections from the lateral geniculate nucleus to the pineal complex of the Mongolian gerbil (Meriones unguiculatus)

The intergeniculate leaflet of the lateral geniculate nucleus is considered to modulate circadian activity rhythms probably mediated by a direct neuronal connection to the suprachiasmatic nucleus. The present study in the gerbil demonstrates, by anterograde tracing with Phaseolus vulgaris-leucoaglutinin (PHA-L), the existence of an additional neuronal projection from a subportion of the lateral geniculate nucleus, involving the intergeniculate leaflet, directly to the pineal gland. PHA-L-immunoreactive nerve fibers originating from perikarya at the injection site were located under the optic tract projecting towards the midsagittal plane. Delicate PHA-L-immunoreactive nerve fibers were observed in the posterior paraventricular thalamic nucleus, precommissural nucleus, olivary pretectal nucleus, anterior and posterior pretectal nuclei, and posterior commissure. Single fibers could be followed from the caudal part of the medial habenular nucleus and the pretectal area into the rostral part of the deep pineal gland. Other fibers continued through the posterior commissure into the contralateral hemisphere to terminate in the same structures as on the ipsilateral side. From the posterior commissure, small bundles of thick fibers entered the deep pineal gland where they arborized among the endocrine cells. A few nerve fibers were observed in the habenular commissure and the pineal stalk, but no fibers were identified in the superficial pineal. This direct geniculo-pineal connection suggests that the pineal gland is directly influenced by the optic system.

Immunocytochemical localization of serotonin and photoreceptor-specific proteins (rod-opsin, S-antigen) in the pineal complex of the river lamprey, Lampetra japonica, with special reference to photoneuroendocrine cells

The pineal complex of the river lamprey, Lampetra japonica, was examined by means of immunocytochemistry with antisera against serotonin, the precursor of melatonin, and two photoreceptor proteins, rod-opsin (the apoprotein of the photopigment rhodopsin) and S-antigen. Serotonin-immunoreactive cells were observed in both the pineal and the parapineal organ. The proximal portion of the pineal organ (atrium) comprised numerous serotonin-immunoreactive cells displaying spherical somata. In the distal end-vesicle of the pineal organ, the serotonin-immunoreactive elements resembled photoreceptors in their size and shape. These cells projecting into the pineal lumen and toward the basal lamina were especially conspicuous in the ventral portion of the end-vesicle. In addition, single serotonin-immunoreactive nerve cells were found in this location. Retinal photoreceptors were never seen to contain immunoreactive serotonin; amacrine cells were the only retinal elements exhibiting serotonin immunoreaction. Strong S-antigen immunoreactivity was found in numerous photoreceptors located in the pineal end-vesicle. In contrast, the S-antigen immunoreactivity was weak in the spherical cells of the atrium. Thus, the pattern of S-antigen immunoreactivity was roughly opposite to that of serotonin. Similar findings were obtained in the parapineal organ. The rod-opsin immunoreaction was restricted to the outer segments of photoreceptors in the pineal end-vesicle and parapineal organ. No rod-opsin++ immunoreactive outer segments occurred in the proximal portion of the atrium. Double immunostaining was employed to investigate whether immunoreactive opsin and serotonin are colocalized in one and the same cell. This approach revealed that (i) most of the rod-opsin-immunoreactive outer segments in the end-vesicle belonged to serotonin-immunonegative photoreceptors; (ii) nearly all serotonin-immunoreactive cells in the end-vesicle bore short rod-opsin-immunoreactive outer segments protruding into the pineal lumen; and (iii) the spherical serotonin-immunoreactive cells in the pineal stalk lacked rod-opsin immunoreaction and an outer segment. These results support the concept that multiple cell lines of the photoreceptor type exist in the pineal complex at an early evolutionary stage.

Direct projections from the anterior and tuberal regions of the lateral hypothalamus to the rostral part of the pineal complex of the rat. An anterograde neuron-tracing study by using Phaseolus vulgaris leucoagglutinin

The anterograde tracer, Phaseolus vulgaris leucoagglutinin, was injected into the lateral hypothalamic division of the Wistar rat. After 7 days of survival the animals were fixed by perfusion, and cryostat sections processed for visualization of the tracer by immunohistochemistry. Injections into the anterior and tuberal regions of the lateral hypothalamic division labeled neurons which projected caudally to the rostral part of the pineal complex, e.g. the deep pineal gland and the pineal stalk. Labeled fibers were in this study not observed in the superficial pineal gland. Due to the direct innercation of the lateral hypothalamic region from the retina and the involvement of this area in circadian rhythmicity, the projections, demonstrated in this study, from the lateral hypothalamic region to the pineal gland are suggested to transmit impulses which modulate the circadian activity of the rat pineal gland.

A direct neural projection from the intergeniculate leaflet of the lateral geniculate nucleus to the deep pineal gland of the rat, demonstrated with Phaseolus vulgaris leucoagglutinin

The anterograde tracer Phaseolus vulgaris leucoagglutinin (PHA-L) was injected into different subregions of the rat lateral geniculate nucleus. After a survival for 5–10 days, the rats were fixed by perfusion with 4% paraformaldehyde, whereafter the brains were cut in a cryostat and the tracer was localized by immunohistochemistry. After deposits of PHA-L involving the intergeniculate leaflet, a high number of PHA-L-immunoreactive fibers were observed to project directky into the posterior commissure. From the posterior commissure, some nerve fibers turned dorsally and entered into the deep pineal gland, a part of the pineal complex located in between the posterior and the habenular commissure. A few PHA-L-immunoreactive fibers were observed in the pineal stalk, but no fibers were detected in the superficial pineal gland. In cases where the injections were placed in the dorsal or ventral subnuclei, no immunoreactive nerve fibers were observed to enter the pineal complex. These results indicate that the intergeniculate leaflet of the lateral geniculate nucleus, a nucleus considered to be involved in circadian rhythmicity, might influence the pineal gland, via a neural projection to the rostral part of the pineal complex.

Quantitative analysis of ?synaptic? ribbon profiles in the pineal complex of male and female Pirbright-White guinea pigs

Previous studies have pointed in the direction of sex differences as well as regional differences in the pineal gland of guinea pigs. In the present investigation these aspects were studied at the electron-microscopic level by quantitating different types of "synaptic" bodies, intrinsic to pinealocytes. The two major types of "synaptic" organelles, ribbons and spherules, did not exhibit regional or sex differences. "Synaptic" structures intermediate in appearance to ribbons and spherules were significantly larger in number in males in the distal region of the pineal gland, compared to females. As previous studies have shown that ribbon and spherule numbers undergo characteristic changes depending on the functional state of the pineal gland, it is concluded that, as far as the "synaptic" organelles are concerned, no clear-cut sex or regional differences appear to exist in the guinea pig pineal gland.

Neural elements of the pineal complex of the frog, rena esculenta, I: Centrally projecting neurons

The pineal complex of anuran amphibians is a directly photosensory organ, encompassing both an extracranial portion, the frontal organ, and an intracranial portion, the pineal organ proper. The projection neurons of the frontal organ respond differentially according to the wavelengths of the light stimuli. The pineal organ, on the other hand, functions mainly as a luminosity meter. Most of its centrally projecting neurons respond to all increases in ambient illumination with decreases in spontaneous firing of action potentials, although some neural units in the pineal organ may respond according to wavelength. This difference in responses to light stimulation may be reflected in the neural organization of the two parts of the pineal complex. In the present study, we have analyzed the morphology of the projection neurons of the frontal and pineal organs of the frog, Rana esculenta, by backfilling of the neurons with horseradish peroxidase through their cut axons. In the pineal organ, several types of centrally projecting neurons were observed: peripherally situated unipolar and multipolar neurons, the dendrites of which extend into a superficial axon plexus that surrounds the pineal epithelium; smaller unipolar, bipolar, or multipolar neurons situated close to the central pineal tract; and radially oriented bipolar neurons, with short dendritic processes oriented towards the lumen of the pineal organ.(ABSTRACT TRUNCATED AT 250 WORDS)

Neural elements in the pineal complex of the frog, Rana esculenta, II: GABA-immunoreactive neurons and FMRFamide-immunoreactive efferent axons

The photosensory pineal complex of anurans comprises an extracranial part, the frontal organ, and an intracranial part, the pineal organ proper. Although the pineal organ functions mainly as a luminosity detector, the frontal organ may monitor the relative proportions of short and intermediate/long wavelengths in the ambient illumination. The major pathway of information processing in the pineal and frontal organs is the photoreceptor to ganglion cell synapse. It is not known whether interneurons form part of the neural circuitry. In the present study, we demonstrate GABA-immunoreactive (GABA-IR) neurons in the pineal and frontal organs of the frog, Rana esculenta. No GABA-IR axons were observed in the pineal nerve between the frontal and pineal organs, or in the pineal tract that connects the pineal complex with the brain. The GABA-IR neurons differed in morphology from centrally projecting neurons visualized by retrograde labeling with horseradish peroxidase. Thus, we suggest that the GABA-IR neurons in the pineal and frontal organs represent local interneurons. Axons of central origin, immunoreactive with a sensitive antiserum against the tetrapeptide Phe-Met-Phe-Arg-NH2 (FMRFamide), were observed in the intracranial portion of the photosensory pineal organ. The immunoreactive axons enter the caudal pole of the pineal organ via the posterior commissure. The largest density of axons was observed in the caudal part, while fewer axons were detected in the rostral portion. The uneven distribution of the FMRFamide-immunoreactive axons may be related to the distribution of different types of intrapineal neurons. FMRFamide-immunoreactive varicose axons were observed in the extracranial frontal organ. A central innervation of the pineal organ, previously known exclusively from amniotes, is probably not per se linked with the evolutionary transition of the pineal organ from a directly photosensory organ to a neuroendocrine organ. It could rather represent a centrifugal input to a sensory system which has been retained when the directly sensory functions have changed, during phylogeny, to neuroendocrine functions.

Electron microscopic analysis of S‐antigen‐ and serotonin‐immuoreactive neural and sensory elements in the photosensory pineal organ of the salmon

Photoreceptor cells in the pineal complex of poikilothermic vertebrates are regarded as homologous with the neuroendocrine pinealocytes in the mammalian pineal organ. They possess an indolamine metabolism, and they contain a number of substances that are immunochemically similar to photo-transduction-related proteins otherwise found in photoreceptors of the lateral eye retina. Using correlative light and electron microscopic pre-embedding immunocytochemistry, we have identified photosensory and neural elements that are immunoreactive with specific antisera against serotonin (5-hydroxy-tryptamine) and the 48 kDa soluble protein S-antigen (arrestin). One type of serotonin-immunoreactive (5HTir) photoreceptor cell was identified. This was characterized by a short basal pole, into which an immunonegative (post-synaptic?) element protruded. Two types of S-antigen-immunoreactive (SAir) photoreceptor cells were observed, one characterized by a short basal pole, similar to that of the 5HTir photoreceptors and the other characterized by a long, extensively branching basal pole. In addition, two types of neurons bearing no morphological specializations typical of photoreceptor cells were SAir: bipolar neurons and multipolar neurons. These were often situated dorsally in the pineal organ. The results indicate an emergence of multiple lines of photoreceptor-derived "pinealocytes" either early in phylogeny, or independently in different taxa. The results are discussed in relation to current theories of pineal evolution.

The pineal complex in Roman high avoidance and Roman low avoidance rats

Previous studies have shown that the pineal gland of Roman high avoidance (RHA/Verh) rats is larger than that of Roman low avoidance rats (RLA/Verh). In the present study measurement of enzyme activities (serotonin-N-acetyl-transferase, hydroxyindole-O-methyltransferase) revealed that pineals of RHA/Verh rats are twice as active in melatonin production than pineals of RLA/Verh rats. Indoleamine content was also higher in RHA/Verh rats, whereas noradrenaline content was the same in both lines. When values were expressed per mg protein, these differences disappeared except for N-acetyl-serotonin and noradrenaline which were higher or lower in RHA/Verh rats, respectively. Both lines had higher serum levels of melatonin during the dark phase than during the light phase. However, RHA/Verh rats had increased serum levels as compared to RLA/Verh rats during both day and night. Morphometric analysis of the deep and superficial part of the pineal complex revealed, that the volumes of both parts are enlarged in RHA/Verh rats. Electron microscopic studies of pineals collected during day- and nighttime showed higher numbers of synaptic ribbons per unit area in pineals of RHA/Verh rats. In pineals collected during June synaptic ribbons displayed a day/night rhythm in RHA/Verh rats only, whereas in glands of both lines collected during November no daily changes were found. These results show that closely related but divergently selected rat lines may differ in pineal ultrastructure and pineal function.

Response to pinealectomy and blinding in vitellogenic female frogs (Rana perezi) subjected to high temperature in autumn

The present experiments were carried out to investigate the effects of pinealectomy and bilateral enucleation on the ovarian activity in Rana perezi frogs maintained in 12-h light--12-h dark photoperiod and 20 +/- 1 degrees C during the vitellogenetic growth in late autumn. These environmental conditions, mainly temperature, induce a gonadal and metabolic response similar to that observed in the natural habitat in summer: a marked ovarian follicular regression, a depletion of the energetic resources from fat bodies and liver, and a minimum in oestradiol circulating levels. This response is partially blocked by pinealectomy and blinding. Protein phosphorus, as an index of vitellogenic proteins, and total ovary lipid content were significantly higher in pinealectomized and blinded frogs with respect to sham-operated animals. Likewise, oestradiol concentrations showed a significant increase during the dark phase of the daily photocycle in pinealectomized and blinded animals. From our results, we can suggest that the arrest of vitellogenesis, the depletion of energetic resources, and the regulation of oestradiol levels induced by the high temperature in Rana perezi frogs can be influenced, at least in part, by the pineal complex and lateral eyes.

Ontogenetic development of S-antigen- and rodopsin immunoreactions in retinal and pineal photoreceptors of Xenopus laevis in relation to the onset of melatonin-dependent color-change mechanisms

In Xenopus laevis Daud., the ontogenetic occurrence of two photoreceptor-specific proteins, S-antigen and rod-opsin, was investigated and correlated to the maturation of the neurohormonal effector system involved in melatonin-dependent color-change mechanisms. Tadpoles ranging from stage 12 to 57 (Nieuwkoop and Faber 1956) were fixed in Zamboni's or Bouin's solution. Frozen or paraffin sections of either total heads or dissected brains and eyes were prepared and treated with highly specific antisera against S-antigen and rod-opsin. In the retina, immunoreactive S-antigen and rod-opsin were first demonstrated in a few centrally located photoreceptors at stage 37/38. Photoreceptors of the peripheral (iridical) portions of the retina gradually became immunoreactive during further development. As in the retina, the first S-antigen-immunoreactive photoreceptors in the pineal complex appeared at stage 37/38. At this and all later stages investigated rod-opsin immunoreactivity was restricted to a few dot-like structures resembling developing pineal outer and inner segments. In most animals rod-opsin immunoreactivity was completely absent from the pineal complex. The analysis of retinal proteins with the immunoblotting technique (Western blot) revealed that the S-antigen antibody bound to a 48-kDa protein and the rod-opsin antibody to a 38-kDa protein. The body lightening reaction was determined with the aid of the melanophore index in larvae fixed in light or darkness, respectively. Aggregation of melanophore melanosomes in darkness (the melatonin-dependent primary chromatic response) first occurred at stage 37/38 when melanophores started to differentiate and became pigmented. These results indicate that in Xenopus laevis (i) the molecular mechanisms of photoreception develop simultaneously in retina and pineal complex; (ii) most pineal photoreceptors differ from retinal rods in that they contain immunoreactive S-antigen but essentially no immunoreactive rod-opsin; and (iii) the differentiation of phototransduction processes coincides with the onset of melatonin-dependent photoneuroendocrine regulation of color-change mechanisms.

Central projections of the pineal complex in the silver lamprey Ichthyomyzon unicuspis.

The central projections of the pineal complex of the silver lamprey Ichthyomyzon unicuspis were studied by injection of horseradish peroxidase. The pineal tract courses caudally along the left side of the habenular commissure, and a few fibers penetrate the brain through the caudalmost portion of this commissure. Most of the fibers, however, continue caudally and enter the brain through the posterior commissure. The pineal tract projects bilaterally to the subcommissural organ, the superficial and periventricular pretectum, the posterior tubercular nucleus, the dorsal and ventral thalamus, the dorsal hypothalamus, the optic tectum, the torus semicircularis, the midbrain tegmentum, and the oculomotor nucleus. A few fibers decussate in the tubercular commissure, but the course of these decusate fibers could not be followed owing to the bilateral nature of the projections. No retrogradely labeled cells were found in the brain. With the exception of the projections to the optic tectum and torus semicircularis, the pineal projections in the silver lamprey are similar to those reported in other anamniote vertebrates.

The neural organization of the pineal complex in the frog: Stratification and regional differences.

Plexiform areas are functionally important units of the pineal complex. The silver impregnation technique by Matsuyama (1972) reveals in the frog that nerve fibres branch and terminate in these areas and that they are presynaptic to nerve cells which are part of the plexiform areas. Histochemically, the nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-diaphorase) method according to Scherer-Singler et al. (1983) indicates that pinealocytes and nerve cells are both stained, a strong stratification of different cell types can be visualized. Plexiform areas of different size exist at the dorsal and ventral surface of the organ; the biggest are located rostrodorsally. Approximately twenty large multipolar nerve cells are localized at the ventral surface of the pineal organ mainly at the rostral and lateral borders. They send long processes mainly in caudal direction. These processes are post-synaptic to the boutons of the endfeet belonging to photosensitive pinealocytes. Processes of several multipolar nerve cells converge to one plexiform area, branch and obviously terminate there. As in horizontal cells the processes of the multipolar cells seem to be capable of both receiving and transmitting signals. On the postsynaptic side of the processes of the multipolar cells are smaller nerve cells juxtaposed to the plexiform areas. They are known as pseudounipolar, unipolar or bipolar cells (Wake et al., 1974); the latter are free of boutons of the pinealocytes, their axons forming the pineal tract. Three different types of photosensitive pinealocytes can be distinguished: a slender, a spherical and a double-cone type. Also, the number of boutons varies in the endfeet: boutons occur alone, in groups of two or three, and in clusters.(ABSTRACT TRUNCATED AT 250 WORDS)

The left habenular nucleus contains a discrete serotonin-immunoreactive subnucleus in the coho salmon ( Oncorhynchus kisutch)

By use of antibodies against serotonin, a discrete subnucleus of putatively serotoninergic neurons was observed in the dorsal subdivision of the left habenular nucleus in the brain of the coho salmon. The subnucleus was observed in salmon of different life-stages: in fingerlings, during smolt transformation, after smolt transformation (in seawater), and after spawning. This finding further emphasizes the close relationship between the pineal organ and the habenular nuclei not only in terms of topographical proximity but also in terms of cytological similarities: cells of the habenular nucleus and the pineal complex have previously been shown to be immunoreactive also with antibodies directed against retinal phototransduction proteins [5]. It also underlines the asymmetric organization of the epithalamic region.