Autonomic nerves terminating on microvessels in the pineal organs of various submammalian vertebrates.

In earlier works we have found that in the mammalian pineal organ, a part of autonomic nerves--generally thought to mediate light information from the retina--form vasomotor endings on smooth muscle cells of vessels. We supposed that they serve the vascular support for circadian and circannual periodic changes in the metabolic activity of the pineal tissue. In the present work, we investigated whether peripheral nerves present in the photoreceptive pineal organs of submammalians form similar terminals on microvessels. In the cyclostome, fish, amphibian, reptile and bird species investigated, autonomic nerves accompany vessels entering the arachnoidal capsule and interfollicular meningeal septa of the pineal organ. The autonomic nerves do not enter the pineal tissue proper but remain in the perivasal meningeal septa isolated by basal lamina. They are composed of unmyelinated and myelinated fibers and form terminals around arterioles, veins and capillaries. The terminals contain synaptic and granular vesicles. Comparing various vertebrates, more perivasal terminals were found in reptiles and birds than in the cyclostome, fish and amphibian pineal organs. Earlier, autonomic nerves of the pineal organs were predominantly investigated in connection with the innervation of pineal tissue. The perivasal terminals found in various submammalians show that a part of the pineal autonomic fibers are vasomotoric in nature, but the vasosensor function of some fibers cannot be excluded. We suppose that the vasomotor regulation of the pineal microvessels in the photosensory submamalian pineal--like in mammals--may serve the vascular support for circadian and circannual periodic changes in the metabolic activity of the pineal tissue. The higher number of perivasal terminals in reptiles and birds may correspond to the higher metabolic activity of the tissues in more differentiated species.

Photopigment coexpression in mammals: comparative and developmental aspects.

In mammals, each cone had been thought to contain only one single type of photopigment. It was not until the early 1990s that photopigment coexpression was reported. In the house mouse, the distribution of color cones shows a characteristic division. Whereas in the upper retinal field the ratio of short wave to middle-to-long wave cones falls in the usual range (1:10), in the ventral retinal field M/L-pigment expression is completely missing. In the transitional zone, numerous dual cones are detectable (spatial coexpression). In other species without retinal division, dual cones appear during development, suggesting that M/L-cones develop from S-cones. Dual elements represent a transitory stage in M/L-cone differentiation that disappear with maturation (transitory coexpression). These two phenomena seem to be mutually exclusive in the species studied so far. In the comparative part of this report the retinal cone distribution of eight rodent species is reported. In two species dual cones appear in adult specimens without retinal division, and dual elements either occupy the dorsal peripheral retina, or make up the entire cone population. This is the first observation proving that all cones of a retina are of dual nature. These species are good models for the study of molecular control of opsin expression and renders them suitable sources of dual cones for investigations on the role and neural connections of this peculiar cone type. In the developmental part, the retinal maturation of other species is examined to test the hypothesis of transitory coexpression. In these species S-pigment expression precedes that of the M/L-pigment, but dual cones are either identified in a small number or they are completely missing from the developing retina. These results exclude a common mechanism for M/L-cone maturation: they either transdifferentiate from S-cones or develop independently.

The system of cerebrospinal fluid-contacting neurons. Its supposed role in the nonsynaptic signal transmission of the brain.

Recent investigations confirm the importance of nonsynaptic signal transmission in several functions of the nervous tissue. Present in various periventricular brain regions of vertebrates, the system of cerebrospinal fluid (CSF)-contacting neurons seems to have a special role in taking up, transforming and emitting nonsynaptic signals mediated by the internal and external CSF and intercellular fluid of the brain. Most of the CSF-contacting nerve cells send dendritic processes into the internal CSF of the brain ventricles or central canal where they form terminals bearing stereocilia and a 9+0-, or 9+2-type cilium. Some of these neurons resemble known sensory cells of chemoreceptor-type, others may be sensitive to the pressure or flow of the CSF, or to the illumination of the brain tissue. The axons of the CSF-contacting neurons transmit information taken up by dendrites and perikarya to synaptic zones of various brain areas. By forming neurohormonal terminals, axons also contact the external CSF space and release various bioactive substances there. Some perikarya send their axons into the internal CSF, and form free endings there, or synapses on intraventricular dendrites, perikarya and/or on the ventricular surface of ependymal cells. Contacting the intercellular space, sensory-type cilia were also demonstrated on nerve cells situated in the brain tissue subependymally or farther away from the ventricles. Among neuronal elements entering the internal CSF-space, the hypothalamic CSF-contacting neurons are present in the magnocellular and parvicellular nuclei and in some circumventricular organs like the paraventricular organ and the vascular sac. The CSF-contacting dendrites of all these areas bear a solitary 9 x 2+0-type cilium and resemble chemoreceptors cytologically. In electrophysiological experiments, the neurons of the paraventricular organ are highly sensitive to the composition of the ventricular CSF. The axons of the CSF-contacting neurons terminate not only in the hypothalamic synaptic zones but also in tel-, mes- and rhombencephalic nuclei and reach the spinal cord as well. The supposed chemical information taken up by the CSF-contacting neurons from the ventricular CSF may influence the function of these areas of the central nervous system. Some nerve cells of the photoreceptor areas form sensory terminals similar to those of the hypothalamic CSF-contacting neurons. Special secondary neurons of the retina and pineal organ contact the retinal photoreceptor space and pineal recess respectively, both cavities being embryologically derived from the 3rd ventricle. The composition of these photoreceptor spaces is important in the photochemical transduction and may modify the activity of the secondary neurons. Septal and preoptic CSF-contacting neurons contain various opsins and other compounds of the phototransduction cascade and represent deep encephalic photoreceptors detecting the illumination of the brain tissue and play a role in the regulation of circadian and reproductive responses to light. The medullo-spinal CSF-contacting neurons present in the oblongate medulla, spinal cord and terminal filum, send their dendrites into the fourth ventricle and central canal. Resembling mechanoreceptors of the lateral line organ, the spinal CSF-contacting neurons may be sensitive to the pressure or flow of the CSF. The axons of these neurons terminate at the external CSF-space of the oblongate medulla and spinal cord and form neurohormonal nerve endings. Based on information taken up from the CSF, a regulatory effect on the production or composition of CSF was supposed for bioactive materials released by these terminals. Most of the axons of the medullospinal CSF-contacting neurons and the magno- and parvicellular neurosecretory nuclei running to neurohemal areas (neurohypophysis, median eminence, terminal lamina, vascular sac and urophysis) do not terminate directly on vessels, instead they form neurohormonal nerve terminals attached by half-desmosomes on the basal lamina of the external and vascular surface of the brain tissue. Therefore, the bioactive materials released from these terminals primarily enter the external CSF and secondarily, by diffusion into vessels and the composition of the external CSF, may have a modulatory effect on the bioactive substances released by the neurohormonal terminals. Contacting the intercellular space, sensory-type cilia were also demonstrated on nerve cells situated subependymally or farther away from the ventricles, among others in the neurosecretory nuclei. Since tight-junctions are lacking between ependymal cells of the ventricular wall, not only CSF-contacting but also subependymal ciliated neurons may be influenced by the actual composition of the CSF besides that of the intercellular fluid of the brain tissue. According to the comparative histological data summarised in this review, the ventricular CSF-contacting neurons represent the phylogenetically oldest component detecting the internal fluid milieu of the brain. The neurohormonal terminals on the external surface of the brain equally represent an ancient form of nonsynaptic signal transmission.

Cerebrospinal fluid contacting neurons in the reduced brain ventricular system of the atlantic hagfish, Myxine glutinosa.

Cerebrospinal fluid (CSF)-contacting neurons are sensory-type cells sending ciliated dendritic process into the CSF. Some of the prosencephalic CSF-contacting neurons of higher vertebrates were postulated to be chemoreceptors detecting the chemical composition of the CSF, other cells may percieve light as "deep encephalic photoreceptors". In our earlier works, CSF-contacting neurons of the mechanoreceptor-type were described around the central canal of the hagfish spinal cord. It was supposed that perceiving the flow of the CSF they are involved in vasoregulatory mechanisms of the nervous tissue. In the present work, we examined the brain ventricular system of the Atlantic hagfish with special reference to the presence and fine structure of CSF-contacting neurons. Myxinoids have an ontogenetically reduced brain ventricular system. In the adult hagfish (Myxine glutinosa) the lumen of the lateral ventricle is closed, the third ventricle has a preoptic-, infundibular and subhabenular part that are not connected to each other. The choroid plexus is absent. The infundibular part of the third ventricle has a medial hypophyseal recess and, more caudally, a paired lateral recess. We found CSF-contacting neurons in the lower part of the third ventricle, in the preoptic and infundibular recess as well as in the lateral infundibular recesses. No CSF-contacting neurons were found in the cerebral aqueduct connecting the subhabenular recess to the fourth ventricle. There is a pineal recess and a well-developed subcommissural organ at the rostral end of the aqueduct. Extending from the caudal part of the fourth ventricle in the medulla to the caudal end of the spinal cord, the central canal has a dorsal and ventral part. Dendrites of CSF-contacting neurons are protruding into the ventral lumen. Corroborating the supposed choroid plexus-like function of the wall of the dorsal central canal, segmental vessels reach a thin area on both sides of the ependymal lining. The perikarya of the CSF-contacting neurons found in the brain ventricles are mainly bipolar and contain granular vesicles of various size. The bulb-like terminal of their ventricular dendrites bears several stereocilia and contains basal bodies as well as mitochondria. Basal bodies emit cilia of the 9+0-type. Cilia may arise from the basal body and accessory basal body as well. The axons run ependymofugally and enter--partially cross--the periventricular synaptic zones. No neurohemal terminals similar to those formed by spinal CSF-contacting neurons of higher vertebrates have been found in the hagfish. We suppose that CSF-contacting neurons transform CSF-mediated non-synaptic information taken up by their ventricular dendrites to synaptic one. A light-sensitive role for some (preoptic?) groups of CSF-contacting neurons cannot be excluded.

Autonomic nerves terminating on smooth muscle cells of vessels in the pineal organ of various mammals.

The significance of autonomic nerves reaching the pincal organ was already investigated in connection to the innervation of pinealocytes and mediating light information from the retina for periodic melatonin secretion. In earlier works we found that some autonomic nerve fibers are not secretomotor but terminate on arteriolar smooth muscle cells in the pineal organ of the mink (Mustela vison). Studying in serial sections the pineal organ of the mink and 15 other mammalian species in the present work, we investigated whether similar axons of vasomotor-type are generally present in the wall of pineal vessels, further, whether they reach the organ via the conarian nerves or via periarterial plexuses. In all species investigated, axons of perivasal nerve bundles were found to form terminal enlargements on the smooth muscle layer of pineal arterioles. The neuromuscular endings contain several synaptic and some granular vesicles. Axon terminals are also present around pineal veins. In serial sections, we found that the so-called conarian autonomic nerves reach the pineal organ alongside pineal veins draining into the great internal cerebral vein. Similar nerves present near arteries of the arachnoid enter the pineal meningeal capsule and septa by arterioles, both perivenous and periarterial nerves form terminals of vasomotor-type. The arteriomotor and venomotor regulation of the tone of the vessels of the pineal organ may serve the vascular support for circadian and circannual periodic changes in metabolic activity of the pineal tissue.

Nonvisual photoreceptors of the deep brain, pineal organs and retina.

The role of the nonvisual photoreception is to synchronise periodic functions of living organisms to the environmental light periods in order to help survival of various species in different biotopes. In vertebrates, the so-called deep brain (septal and hypothalamic) photoreceptors, the pineal organs (pineal- and parapineal organs, frontal- and parietal eye) and the retina (of the "lateral" eye) are involved in the light-based entrain of endogenous circadian clocks present in various organs. In humans, photoperiodicity was studied in connection with sleep disturbances in shift work, seasonal depression, and in jet-lag of transmeridional travellers. In the present review, experimental and molecular aspects are discussed, focusing on the histological and histochemical basis of the function of nonvisual photoreceptors. We also offer a view about functional changes of these photoreceptors during pre- and postnatal development as well as about its possible evolution. Our scope in some points is different from the generally accepted views on the nonvisual photoreceptive systems. The deep brain photoreceptors are hypothalamic and septal nuclei of the periventricular cerebrospinal fluid (CSF)-contacting neuronal system. Already present in the lancelet and representing the most ancient type of vertebrate nerve cells ("protoneurons"), CSF-contacting neurons are sensory-type cells sitting in the wall of the brain ventricles that send a ciliated dendritic process into the CSF. Various opsins and other members of the phototransduction cascade have been demonstrated in telencephalic and hypothalamic groups of these neurons. In all species examined so far, deep brain photoreceptors play a role in the circadian and circannual regulation of periodic functions. Mainly called pineal "glands" in the last decades, the pineal organs actually represent a differentiated form of encephalic photoreceptors. Supposed to be intra- and extracranially outgrown groups of deep brain photoreceptors, pineal organs also contain neurons and glial elements. Extracranial pineal organs of submammalians are cone-dominated photoreceptors sensitive to different wavelengths of light, while intracranial pineal organs predominantly contain rod-like photoreceptor cells and thus scotopic light receptors. Vitamin B-based light-sensitive cryptochromes localized immunocytochemically in some pineal cells may take part in both the photoreception and the pacemaker function of the pineal organ. In spite of expressing phototransduction cascade molecules and forming outer segment-like cilia in some species, the mammalian pineal is considered by most of the authors as a light-insensitive organ. Expression of phototransduction cascade molecules, predominantly in young animals, is a photoreceptor-like characteristic of pinealocytes in higher vertebrates that may contribute to a light-percepting task in the perinatal entrainment of rhythmic functions. In adult mammals, adrenergic nerves--mediating daily fluctuation of sympathetic activity rather than retinal light information as generally supposed--may sustain circadian periodicity already entrained by light perinatally. Altogether three phases were supposed to exist in pineal entrainment of internal pacemakers: an embryological synchronization by light and in viviparous vertebrates by maternal effects (1); a light-based, postnatal entrainment (2); and in adults, a maintenance of periodicity by daily sympathetic rhythm of the hypothalamus. In addition to its visual function, the lateral eye retina performs a nonvisual task. Nonvisual retinal light perception primarily entrains genetically-determined periodicity, such as rod-cone dominance, EEG rhythms or retinomotor movements. It also influences the suprachiasmatic nucleus, the primary pacemaker of the brain. As neither rods nor cones seem to represent the nonvisual retinal photoreceptors, the presence of additional photoreceptors has been supposed. Cryptochrome 1, a photosensitive molecule identified in retinal nerve cells and in a subpopulation of retinal photoreceptors, is a good candidate for the nonvisual photoreceptor molecule as well as for a member of pacemaker molecules in the retina. When comparing various visual and nonvisual photoreceptors, transitory, "semi visual" (directional) light-perceptive cells can be detected among them, such as those in the parietal eye of reptiles. Measuring diffuse light intensity of the environment, semivisual photoreceptors also possess some directional light perceptive capacity aided by complementary lens-like structures, and screening pigment cells. Semivisual photoreception in aquatic animals may serve for identifying environmental areas of suitable illumination, or in poikilotermic terrestrial species for measuring direct solar irradiation for thermoregulation. As directional photoreceptors were identified among nonvisual light perceptive cells in the lancelet, but eyes are lacking, an early appearance of semivisual function, prior to a visual one (nonvisual --> semivisual --> visual?) in the vertebrate evolution was supposed.

Pineal organ-like organization of the retina in megachiroptean bats.

Phylogenetically originated from photoreceptive structures, the pineal organ adapts the organism to circadian and circannual light periodicity of the environment, while the retina develops to a light-based locator. Bats have a nocturnal life and an echolocator orientation presumably modifying the task of photoreception. Looking for morphological basis of the special functions, in the present work we compared the fine structure and immunocytochemistry of the retina and pineal organ in micro- and megacrochiroptean bats. We found that there is a high similarity between the retina and pineal organ in megachiropterans when compared to other species investigated so far. Besides of photoreceptor derived pinealocytes, the pineal organ of both micro- and megachiropterans contain intrapineal neurons and/or ganglionic cells as well as glial cells. Like spherules and pedicles of retinal photoreceptors, axon-type processes of pinealocytes form synaptic ribbon containig terminals. Similar to retinal photoreceptors and neurons, pinealocytes and pineal neurons contain immunoreactive glutamate and aspartate. In addition, excitatory amino acids accumulate in the pineal neurohormonal endings and might have a role in the hormonal (serotonin?) release of the organ. Concerning the structure of the retina the highest similarity to the organization of the pineal organ was found in the megachiroptean fruit eating bats Cynopterus sphinx and Rusettus niloticus. The retina of these species forms folds and crypts in its photoreceptor layer. This organization is similar to the folds of the pineal wall successively developed during evolution. Since a folded photoreceptor layer is not viable for a photolocator screen in decoding two-dimensional images, we suppose that this peculiar organization of the megachiropteran retina is connected to a "pineal-like" photometer task of the eye needed by these species active at night.

Comparative ultrastructure and cytochemistry of the avian pineal organ.

The breeding of birds is expected to solve problems of nourishment for the growing human population. The function of the pineal organ synchronizing sexual activity and environmental light periods is important for successful reproduction. Comparative morphology of the avian pineal completes data furnished by experiments on some frequently used laboratory animals about the functional organization of the organ. According to comparative histological data, the pineal of vertebrates is originally a double organ (the "third" and the "fourth eye"). One of them often lies extracranially, perceiving direct solar radiation, and the other, located intracranially, is supposed to measure diffuse brightness of the environment. Birds have only a single pineal, presumably originating from the intracranial pineal of lower vertebrates. Developing from the epithalamus, the avian pineal organ histologically seems not to be a simple gland ("pineal gland") but a complex part of the brain composed of various pinealocytes and neurons that are embedded in an ependymal/glial network. In contrast to organs of "directional view" that develop large photoreceptor outer segments (retina, parietal pineal eye of reptiles) in order to decode two-dimensional images of the environment, the "densitometer"-like pineal organ seems to increase their photoreceptor membrane content by multiplying the number of photoreceptor perikarya and developing follicle-like foldings of its wall during evolution ("folded retina"). Photoreceptor membranes of avian pinealocytes can be stained by antibodies against various photoreceptor-specific compounds, among others, opsins, including pineal opsins. Photoreceptors immunoreacting with antibodies to chicken pinopsin were also found in the reptilian pineal organ. Similar to cones and rods representing the first neurons of the retina in the lateral eye, pinealocytes of birds possess an axonal effector process which terminates on the vascular surface of the organ as a neurohormonal ending, or forms ribbon-containing synapses on pineal neurons. Serotonin is detectable immunocytochemically on the granular vesicles accumulated in neurohormonal terminals. Pinealocytic perikarya and axon terminals also bind immunocytochemically recognizable excitatory amino acids. Peripheral autonomic fibers entering the pineal organ through its meningeal cover terminate near blood vessels. Being vasomotor fibers, they presumably regulate the blood supply of the pineal tissue according to the different levels of light-dependent pineal cell activity.

Comparative histology of pineal calcification.

The pineal organ (pineal gland, epiphysis cerebri) contains several calcified concretions called "brain sand" or acervuli (corpora arenacea). These concretions are conspicuous with imaging techniques and provide a useful landmark for orientation in the diagnosis of intracranial diseases. Predominantly composed of calcium and magnesium salts, corpora arenacea are numerous in old patients. In smaller number they can be present in children as well. The degree of calcification was associated to various diseases. However, the presence of calcified concretions seems not to reflect a specific pathological state. Corpora arenacea occur not only in the actual pineal tissue but also in the leptomeninges, in the habenular commissure and in the choroid plexus. Studies with the potassium pyroantimonate (PPA) method on the ultrastructural localization of free calcium ions in the human pineal, revealed the presence of calcium alongside the cell membranes, a finding that underlines the importance of membrane functions in the production of calcium deposits. Intrapineal corpora arenacea are characterized by a surface with globular structures. Meningeal acervuli that are present in the arachnoid cover of the organ, differ in structure from intrapineal ones and show a prominent concentric lamination of alternating dark and light lines. The electron-lucent lines contain more calcium than the dark ones. There is a correlation between the age of the subject and the number of layers in the largest acervuli. This suggests that the formation of these layers is connected to circannual changes in the calcium level of the organ. The histological organization of the human pineal is basically the same as that of mammalian experimental animals. Pineal concretions present in mammalian animal species are mainly of the meningeal type. Meningeal cells around acervuli contain active cytoplasmic organelles and exhibit alkaline phosphatase reaction in the rat and mink, an indication of a presumable osteoblast-like activity. Using Kossa's method for the staining of calcium deposits, a higher calcium concentration was detected in the rat pineal than in the surrounding brain tissue. Since in parathyroidectomised rats calcified deposits are larger and more numerous than in controls, the regulation of the production of acervuli by the parathyroid gland has also been postulated. In most of submammalian species, the pineal organs (pineal-, parapineal organ, frontal organ, parietal eye) are photoreceptive and organized similarly to the retina. Acervuli were found in the pineal of some birds. The pineal organs of lower vertebrates (fish, amphibians, reptiles) exhibit a high calcium content by ultrastructural calcium histochemistry (PPA-method). However, concrements are not formed. The accumulation of Ca2+ seems to depend on the receptor function of the organ. Comparing pineal and retinal photoreceptors in the frog, the photoreceptor outer segments of pinealocytes as well as retinal cones and rods show a large amount of Capyroantimonate deposits. In dark adapted animals calcium ions are present in both sides of the photoreceptor membranes of the outer segment, whereas calcium is shifted extra-cellularly following light adaptation. Overviewing the data available about the pineal calcification, we can conclude that a multifactorial mechanism may be responsible for the calcification. The pineal of higher vertebrates is not just a simple endocrine gland, rather, its histological organization resembles a folded retina having both hormonal and neural efferentation. Mammalian pinealocytes preserve several characteristics of submammalian receptor cells and accumulate free Ca2+ on their membranes (1). In the thin walled retina and in the similarly organized pineal of submammalian species, the diffusion of extracellular calcium is probably easy and there is a lesser tendency to form concrements. (ABSTRACT TRUNCATED)

Actual problems of the cerebrospinal fluid-contacting neurons.

Cerebrospinal fluid (CSF)-contacting neurons form a part of the circumventricular organs of the central nervous system. Represented by different cytologic types and located in different regions, they constitute a CSF-contacting neuronal system, the most central periventricular ring of neurons in the brain organized concentrically according to our concept. Because the central nervous system of deuterostomian echinoderm starfishes and the prochordate lancelet is composed mainly of CSF-contacting-like neurons, we hypothesize that this cell type represents ancient cells, or protoneurons, in the vertebrate brain. Neurons may contact the ventricular CSF via their dendrites, axons, or perikarya. Most of the CSF-contacting nerve cells send their dendritic processes into the ventricular cavity, where they form ciliated terminals. These ciliated endings resemble those of known sensory cells. By means of axons, the CSF-contacting neurons also may contact the external CSF space, where the axons form terminals of neurohormonal type similar to those known in the neurohemal areas. The most simple CSF-contacting neurons of vertebrates are present in the terminal filum, spinal cord, and oblongate medulla. The dendritic pole of these medullospinal CSF-contacting neurons terminates with an enlargement bearing many stereocilia in the central canal. These cells are also supplied with a 9 x 2 + 2 kinocilium that may contact Reissner's fiber, the condensed secretory material of the subcommissural organ. The Reissner's fiber floating freely in the CSF leaves the central canal at the caudal open end of the terminal filum in lower vertebrates, and open communication is thus established between internal CSF and the surrounding tissue spaces. Resembling mechanoreceptors cytologically, the spinal CSF-contacting neurons send their axons to the outer surface of the spinal cord to form neurosecretory-type terminals. They also send collaterals to local neurons and to higher spinal segments. In the hypothalamic part of the diencephalon, neurons of two circumventricular organs, the paraventricular organ and the vascular sac, of the magnocellular neurosecretory nuclei and several parvocellular nuclei, form CSF-contacting dendritic terminals. A CSF-contacting neuronal area also was found in the telencephalon. The CSF-contacting dendrites of all these areas bear solitary 9 x 2 + 0 cilia and resemble chemoreceptors and developing photoreceptors cytologically. In electrophysiological experiments, the neurons of the paraventricular organ are highly sensitive to the composition of the ventricular CSF. The axons of the CSF-contacting neurons of the paraventricular organ and hypothalamic nuclei terminate in hypothalamic synaptic zones, and those of magno- and parvocellular neurosecretory nuclei also form neurohormonal terminals in the median eminence and neurohypophysis. The axons of the CSF-contacting neurons of the vascular sac run in the nervus and tractus sacci vasculosi to the nucleus (ganglion) sacci vasculosi. Some hypothalamic CSF-contacting neurons contain immunoreactive opsin and are candidates to represent the "deep encephalic photoreceptors." In the newt, cells derived from the subependymal layer develop photoreceptor outer segments protruding to the lumen of the infundibular lobe under experimental conditions. Retinal and pineal photoreceptors and some of their secondary neurons possess common cytologic features with CSF-contacting neurons. They contact the retinal photoreceptor space and pineal recess, respectively, both cavities being derived from the third ventricle. In addition to ciliated dendritic terminals, there are intraventricular axons and neuronal perikarya contacting the CSF. Part of the CSF-contacting axons are serotoninergic; their perikarya are situated in the raphe nuclei. Intraventricular axons innervate the CSF-contacting dendrites, intraventricular nerve cells, and/or the ventricular surface of the ependyma. (ABSTRACT TRUNCATED)

The pineal organ as a folded retina: immunocytochemical localization of opsins.

The most simple pineal complex (the pineal and parapineal organs of lampreys), consists of saccular evaginations of the diencephalic roof, and has a retina-like structure containing photoreceptor cells and secondary neurons. In more differentiated vertebrates, the successive folding of the pineal wall multiplies the cells and reduces the lumen of the organ, but the pattern of the histological organization remains similar to that of lampreys; therefore, we consider the histological structure of the pineal organ of higher vertebrates as a 'folded retina'. The cell membrane of several pineal photoreceptor outer-segments of vertebrates immunoreact with anti-retinal opsin antibodies supporting the view of retina-like organization of the pineal. Some other pineal outer segments do not react with retinal anti-opsin antibodies, a result suggesting the presence of special pineal photopigments in different types of pinealocytes that obviously developed during evolution. The chicken pinopsin, detected in the last years, may represent one of these unknown photopigments. Using antibodies against chicken pinopsin, we compared the immunoreactivity of different photoreceptors of the pineal organs from cyclostomes to birds at the light and electron microscopic levels. We found pinopsin immunoreaction on all pinealocytes of birds and on the rhodopsin-negative large reptilian pinealocytes. As the pinopsin has an absorption maximum at 470 nm, these avian and reptilian immunoreactive pinealocytes can be regarded as green-blue light-sensitive photoreceptors. Only a weak immunoreaction was observed on the frog and fish pinealocytes and no reaction was seen in cyclostomes and in the frontal organ of reptiles. Some photoreceptors of the retina of various species also reacted the pinopsin antibodies, therefore, pinopsin must have certain sequential similarity to individual retinal opsins of some vertebrates.

Immunoreactive excitatory amino acids in the parietal eye of lizards, a comparison with the pineal organ and retina.

The fine structure of the organ and the localization of the excitatory amino acids glutamate and aspartate were studied in the parietal eye of lizards by postembedding immunoelectron microscopy. The parietal eye contains cone photoreceptor cells, secondary neurons, and ependymal and lens cells. The photoreceptors form long inner and outer segments, some of them being paired as "twin-photoreceptors" by zonulae adherentes. Perikarya of neurons bear sensory cilia (containing 9x2+0 pairs of tubules) extending into the intercellular space. No neurohormonal terminals are present in the parietal eye. A higher immunoreactivity to glutamate than to aspartate is found in the photoreceptors and in the secondary neurons of the parietal eye. Glutamate immunogold labeling is more intense in the axonal processes of photoreceptors and neurons and in most of the nerve fibers of the parietal nerve running to the brain stem. Weak aspartate and glutamate immunoreactivity can be detected in the ependymal and lens cells. A similar distribution of immunoreactive amino acids is found in the photoreceptors, secondary neurons, and ependymal glial elements of the pineal organ, and retina of the lateral eye of the same animals. Immunoreactive glutamate accumulates in the axons of photoreceptors and secondary neurons of the parietal eye suggesting that this excitatory amino acid acts as a synaptic mediator in the neural efferentation of the organ. Thus, the efferent light-conducting pathway of the parietal organ is similar to that of the pineal organ and lateral eye retina. As the Mullerian cells of the retina, the ependymal and lens cells of the parietal eye and the ependymal-glial cells of the pineal organ may play a role in the metabolism and/or elimination of excitatory amino acids released by photoreceptors.

Mediator substances of the pineal neuronal network of mammals.

In addition to receptor-type pinealocytes, the mammalian pineal organ contains small and large neurons and ependymal/glial cells as well. Axons of pinealocytes form synaptic ribbon-containing axo-dendritic synapses on large secondary pineal neurons and/or terminate as neurohormonal endings on the basal lamina of the vascular surface of the organ. The small pineal neurons were found to be gamma-aminobutyric acid (GABA)-immunoreactive, while large secondary neurons and pinealocytes contained immunoreactive amino acids (glutamate and aspartate). Glutamate accumulated presynaptically in pinealocytic axon terminals on large secondary neurons and in the axons of these neurons. Glutamate immunoreactive axons of pineal neurons were traced via the pineal tract to the habenular nucleus. Axons containing granular vesicles and coming from extrapineal perikarya are glutamate immunoreactive as well. Aspartate and GABA are also present in some of the myelinated axons, supposedly pinealopetal in the pineal tract.

Immunoreactive pinopsin in pineal and retinal photoreceptors of various vertebrates.

Pinopsin is a pineal specific opsin newly identified in the pineal of birds which has an absorption maximum at 470 nm. As the opsin content of photoreceptors in the pineal complex of several species is not yet known, in the present work, we studied their pinopsin immunoreactivity in various vertebrates from cyclostomes to mammals. We also compared the immunoreactivity of pineal photoreceptors to that of retinal cones and rods of each animal. For the immunocytochemistry, we raised antibodies in rabbits against a 14 amino acids containing part of the chicken pinopsin molecule. The immunoreaction was performed at the electron microscopic level. The pineal organs show a great diversity in vertebrates: there is a pineal organ present from cyclostomes to mammals, in addition, there is a parapineal organ in cyclostomes and fishes, a frontal organ in frogs and a parietal eye in several reptiles. We detected a strong pinopsin immunoreaction on most of the pinealocytes of birds and on the large photoreceptor-type of the pineal of reptiles. Rod-type photoreceptors of the avian retina and a cone of the reptile retina was immunoreactive as well. According to the known absorption maximum of pinopsin, the immunoreactivity may indicate a green-blue light-sensitivity for these photoreceptors. The immunoreactivity was less pronounced or absent in mammals as well as in less differentiated species. The pineal organ of snakes and the parietal eye of reptiles equally failed to exhibit pinopsin immunoreactive photoreceptors, presumably, due to the absence of green-blue light-sensitive photoreceptors of pinopsin-type in these species.

Similar fine structural localization of immunoreactive glutamate in the frog pineal complex and retina.

The distribution of immunoreactive glutamate was compared in the pineal complex (pineal and frontal organs) and retina of frogs (Rana esculenta, R. arvalis, R. ridibunda, R. catesbeiana, Bufo viridis, Bombinator igneus) by postembedding immuno-electron microscopy. Similar to retinal photoreceptors (rods and cones), bipolars and ganglion cells, the rod- and cone-like photoreceptors and the neurons of the pineal and frontal organs exhibited glutamate immunoreactivity. Synaptic terminals of photoreceptor cells on secondary neurons of the pineal complex and retina were strongly immunoreactive. The pineal tract and the fibers of the frontal nerve also displayed glutamate immunoreactivity. There was no essential difference in the immunoreactivity of the retinal and pineal elements among the species studied. The similar histology of the pineal complex and retina of the frog and the high correlation of their binding sites of antiglutamate immunosera allow us to assume that glutamate performs a similar role in the pineal complex as is already known for the retina. The high immunoreactivity of the presynaptic region of pinealocytic processes and axons of secondary neurons suggests the role of a neurotransmitter for this excitatory amino acid in the efferent pathways of the pineal complex.

Similar localization of immunoreactive glutamate and aspartate in the pineal organ and retina of various nonmammalian vertebrates.

The localization of immunoreactive glutamate and aspartate was compared in the pineal organ and retina of various vertebrates (Raja clavata, Carassius auratus, Salvelinus alpinus, Triturus vulgaris, Triturus cristatus, Lacerta muralis, Lacerta agilis, Lacerta viridis, Columbia livia and white leghorn chicken) by postembedding immunoelectron microscopy. Immunoreaction of both excitatory amino acids was detected in the pinealocytes in a localization similar to that of retinal photoreceptors. The reaction was intense in the axonal processes of pinealocytes as well as retinal rods and cones, further in their terminals on secondary pineal and retinal neurons. Subsequent immunoreaction on the same section showed a colocalization of glutamate and aspartate. The accumulation of these amino acids in the presynaptic part of pinealocytes suggests that they act as synaptic mediators in the neural efferentation of the pineal organ. In reptiles and birds where the hormonal efferentation of the pineal is well developed, glutamate and aspartate was also found to be accumulated in neuroendocrine terminals of pinealocytes. Therefore, glutamate and aspartate may have a role in both the hormonal and neural efferentation of the pineal organ.

Development of the photoreceptor outer segment-like cilia of the CSF-contacting pinealocytes of the ferret (Putorius furo).

The postnatal development of the pineal organ of the ferret (Putorius furo) was investigated electron-microscopically with special interest given to the cerebrospinal fluid (CSF)-contacting pinealocytes and their large, vesiculated cilia. In the pineal of the newborn ferrets, there is a lumen--a pineal ventricle--which is a diverticle of the third ventricle of the diencephalon. The luminal surface of the pineal is bordered by ependymal cells and CSF-contacting pinealocytes. A sensory, 9 x 2 + 0 type cilium arises from the free surface of the pinealocytes and thickens in the first week. There are mitotic figures in the wall of the pineal ventricle, being reduced to a pineal recess during the second and third postnatal week. In two week-old animals, vesicles appear in the cilia of the pinealocytes. The vesicles may form rows and fill the enlarged cilium at the third week. Near the basal bodies, a proximal connecting piece remains narrow and free of vesicles. In older animals, there are multivesicular and dense bodies in the pineal cilia. The reduction of the pineal ventricle closes the CSF-contacting cilia in the intercellular spaces. Axon-like processes of pinealocytes form synaptic ribbon-containing terminals on secondary pineal neurons. Axons of pineal neurons enter the fiber bundles of the pineal tract running to the habenular nuclei. All these structures do not differ from the light conducting pathway of the submammalian pineals. The ultrastructure of the cilia investigated resembles that of the developing outer segments of the retina and represents a preserved light perceiving structure of the mammalian pinealocytes. Further studies are necessary to elucidate whether the early differentiation of the cilia and synapses indicates a timing of the circadian light rhythmicity in young ferrets by direct pineal photosensitivity.