Pathology of the hippocampus in experimental feline infantile hydrocephalus.

Hydrocephalus is responsible for many pediatric neurological deficits presumed to be caused by neocortical pathophysiology. Relatively little is known about the role of non-neocortical CNS structures in this condition. In the present work experimental infantile hydrocephalus produced by intracisternal kaolin injection was studied in a neonate kitten model. The hippocampal formation was processed for electron microscopy, and the neuropil of the CA3 region was examined in untreated, severely hydrocephalic and age-matched normal animals. Both macroscopically and microscopically the thickness of the hippocampus was not decreased. Hippocampal pyramidal neurons were found in varying stages of cytoplasmic densification, and dendritic and axonal processes exhibited hydropic cellular deterioration. The number of synaptic contacts was decreased. However, there was no indication of edematous extracellular space and the ependymal covering of the hippocampus was intact. The macroscopic structural integrity of the hippocampus, as well as the dendritic, axonal and synaptic alterations, suggest that the dark pyramidal neurons are the result of deafferentation, which may have profound effects on learning and memory.

The microglial response to progressive hydrocephalus in a model of inherited aqueductal stenosis.

Although gliosis has been reported to be a common and persistent feature in the white matter of hydrocephalic brains, no studies have identified the cell types that characterize this response. Therefore, the present study has employed histochemical methods to evaluate microglial cells in the brains of infant rats with inherited hydrocephalus. This strain of rats acquires hydrocephalus during late fetal stages due to aqueductal stenosis. Tissue from the sensorimotor and auditory cortices of 12- and 21-day-old hydrocephalic and normal H-Tx rats was processed and stained for the lectin microglial marker Griffonia simplicifolia (GSA-IB4). During the progression of hydrocephalus, GSA-positive cells exhibited three changes: (1) Cytologically, the cell bodies were enlarged, and their processes were thicker, longer and more numerous. These changes were most notable in the gray matter. (2) The packing density of GSA-positive cells was either increased or decreased, depending on the age of the animal and the severity of hydrocephalus. (3) Localized clusters of GSA-positive cells were conspicuous in the white matter of 12-day animals with mild hydrocephalus, and in the gray matter of 21-day animals with severe hydrocephalus. These results indicate that the microglial response is initiated during intermediate stages of hydrocephalus, and is not restricted to the periventricular white matter. These changes may signal other pathophysiologic events in the hydrocephalic brain, and demonstrate that microglia constitute one important element in the gliosis that accompanies hydrocephalus.

The microstructure of cortical neuropil before and after decompression in experimental infantile hydrocephalus.

Hydrocephalus is a common clinical disorder and responsible for many pediatric neurological deficits. Relatively little is known about the cellular mechanisms of this disorder and less is known about reconstitution of connectivity following ventricular shunt procedures. In the present studies experimental infantile hydrocephalus produced by kaolin injection was studied in a neonate kitten model. The neuropil of the cerebral cortex was examined in hydrocephalic animals and animals which received a ventriculoperitoneal shunt to reduce ventriculomegaly. The brains were processed for Golgi silver impregnation and electron microscopy to study the detailed dendritic and synaptic architecture. The periventricular region of the hydrocephalic animals exhibited increased extracellular space and signs of neuronal degeneration. Components of the deep neuropil (laminae V-VI) were in disarray and surrounded by edematous extracellular spaces. The superficial neuropil (laminae I-IV), in contrast, appeared intact, but detailed examination showed indications of dendritic degeneration. Shunt procedures successfully restored the cortical mantle to near normal thickness. However, Golgi light microscopy and electron microscopy revealed that dendritic appendage morphology was altered. The results are discussed in regard to development of neuronal connectivity following shunt procedures.

Biogenic amine localization in cardiac ganglion intrinsic neurons: electron microscopic histochemistry of SIF cells.

The parasympathetic cardiac ganglion in the mudpuppy, N. maculosus, contains postganglionic nerve cells and intrinsic neurons, many of which are small intensely fluorescent (SIF) cells. Several bioactive substances have been localized in the intrinsic nerve cells which may have integrative effects at synapses within the ganglion. Ganglionic intrinsic neurons can be identified electron microscopically by the presence of numerous cytoplasmic granular vesicles 80-120 nm in diameter. Throughout the ganglion there are bundles of unmyelinated fibers some of which are filled with granular and agranular vesicles and axosomatic terminals with similar vesicles synapsing on principal parasympathetic nerve cells. To understand the aminergic contribution to ganglionic synaptic circuitry the chromaffin reaction was used. The intrinsic neurons (i.e., SIF cells) were readily identified by their characteristic intracellular granule population. All intrinsic nerve cells identified showed granules which were positively labelled by the chromaffin reaction. Granular vesicles in synaptic profiles on principal cells (P cells) were also labelled indicating a direct aminergic synaptic innervation to these cells. The cell bodies of intrinsic neurons, ensheathed with supportive glial-like cellular processes, rarely received synapses. Elemental microanalysis was used to verify the chromium content of the electron dense product within the granular vesicles. These studies demonstrated direct aminergic synaptic input to at least a subpopulation of principal parasympathetic cells in the cardiac ganglion of mudpuppy.

Rete mirabile medullae: vascular specialization of the avian brain stem.

A circumscribed vascular plexus is located between the conjoint proximal ganglion of cranial nerves IX-X and the lateral surface of the avian brain stem. Gross, light, and electron microscopic analyses reveal that this structure is a rete mirabile. This rete is unique in that it resides in the subarachnoid space where it is bathed by cerebrospinal fluid. Microscopic and ultrastructural studies document that the rete is composed of arterioles with wide luminal diameters, marginal folds project from the endothelial cells into the lumen of the blood vessel. The endothelial cells contain numerous pinocytotic vesicles. The underlying basal lamina is penetrated by myoendothelial junctions. One to three layers of smooth muscle cells comprise the tunica media. Bundles of unmyelinated nerve fibers course through the loose connective tissue encompassing the blood vessels.

Distribution of serotonin in the caudal neurosecretory complex. A light and electron microscopic study.

The caudal neurosecretory complex (CNc) of poecilids has previously been shown to receive serotonergic inputs. In the present study, immunohistochemical techniques were applied at the light and electron microscopic levels to characterize serotonergic terminals in the neuroendocrine nucleus. A dense plexus of varicose fibers observed in the rostral CNc neuropil was absent in the spinal cords of deafferented fish, indicating that the origin of this input was extranuclear. Ultrastructural study revealed no direct contacts between labeled structures and neuroendocrine cells. Non-synaptic terminals (varicosities) were the predominantly labeled structures in the neuropil. Synaptic terminals were observed on cellular and axonal targets in the CNc. Small cells containing 70 nm dense-core vesicles received serotonergic input on their perikarya. Labeled synapses were also found on unlabeled axon terminals which made axo-axonal synapses on neuroendocrine processes. Non-synaptic terminals may be responsible for a variety of serotonin-mediated effects in the CNc. Synaptic interactions with local catecholaminergic and afferent cholinergic inputs to the CNc are likely.

Brainstem location of serotonin neurons projecting to the caudal neurosecretory complex.

Serotonergic fibers in the caudal neurosecretory complex (CNc) of poeciliids originate from neurons within, and extrinsic to this spinal cord nucleus. In the present study, retrograde tracing and immunofluorescence techniques were combined to localize extrinsic serotonergic projection neurons. The entire spinal cord and brain were sectioned after Fast Blue (FB) or horseradish peroxidase (HRP) was implanted in the CNc. No HRP or FB filled neurons were found in the spinal cord. Retrogradely filled neurons were found bilaterally in dorsolateral and ventromedial reticular nuclei, and the dorsal midbrain tegmentum. Fusiform cells in the medullary fasciculus longitudinalis medialis filled with FB but not HRP. Serotonin immunopositive neurons were found surrounding the third ventricle, in the raphe and in medullary reticular nuclei. Double labelled neurons in the medial reticular nucleus were determined to be the source of serotonergic projections to the CNc. Reticular projection nuclei are strategically situated to receive visceral sensory input from rhombencephalic cranial nerves. These putative pathways may provide an anatomical substrate by which visceral sensory information is transmitted to the CNc.

Terminal processes of serotonin neurons in the caudal spinal cord of the molly, Poecilia latipinna, project to the leptomeninges and urophysis.

The caudal neurosecretory complex of poeciliids has previously been shown to be innervated by extranuclear and intrinsic serotonergic projections. In the present study, immunohistochemical techniques were used to characterize fibers originating from serotonin neurons intrinsic to the caudal spinal cord. Bipolar and multipolar neurons were oriented ventromedially, and contained numerous large granular vesicles. Three types of serotonergic fibers were distinguished based on their distribution and morphology. Intrinsic Type-A fibers branched into varicose segments near the ventrolateral surface of the spinal cord and contacted the basal lamina beneath the leptomeninges. Type-B fibers coursed longitudinally to enter the urophysis, where they diverged and terminated around fenestrated capillaries. Labelled vesicles in Type-A and Type-B terminals were the same size as those in labelled cells and in unlabelled neurosecretory terminals in the urophysis. Type-C small varicose fibers branched within the neuropil of the caudal neurosecretory complex. Serotonin may be secreted into the submeningeal cerebrospinal fluid, the urophysis, and the caudal vein by Type-A and Type-B fibers, whereas, Type-C fibers may be processes of serotonergic interneurons in the neuroendocrine nucleus. The possibility that urotensins I and II or arginine vasotocin were colocalized in the processes of the intrinsic serotonin neurons was investigated immunohistochemically. The negative results of these experiments suggest that serotonin-containing neurons may represent a neurochemically distinct subpopulation in the caudal neurosecretory complex.

Monoamines in the caudal neurosecretory complex: biochemistry and immunohistochemistry.

Monoaminergic inputs to the caudal neurosecretory complex (CNc) of Poecilia latipinna have been identified using histofluorescence and immunohistochemical techniques. The present study was undertaken to identify specific monoamines and determine the relative contribution of indolamines and catecholamines in supraspinal and intrinsic innervation of the nucleus. The CNc was deafferented by transecting the spinal cord rostral to the CNc. Ten days subsequently, CNc of spinal-transected and control fish were processed for either biochemical or immunohistochemical analysis. Norepinephrine and serotonin were detected in pooled samples of control CNc. Following deafferentation, the content of both monoamines was diminished. Using immunohistochemical labeling for serotonin or for the catecholamine-synthesizing enzymes, tyrosine hydroxylase (TH) or dopamine-beta-hydroxylase (DBH), the number of monoamine fibers was decreased in deafferented CNc compared to control. A substantial serotonergic innervation remains after deafferentation, as evidenced by serotonin-positive neurons and heavy, varicose fibers. Occasional TH/DBH-positive cells and fibers remain after deafferentation. These data suggest that both norepinephrine and serotonin are associated with descending supraspinal projections, while serotonin predominates as the intrinsic monoamine.

Telencephalic projections to the goldfish hypothalamus: an anterograde degeneration study.

In this study, large areas of goldfish telencephalon were ablated including rostral nucleus preopticus periventriculare (rNPP), and degenerating axons were traced by a modified Fink and Heimer procedure. The lesioning procedure ablated large regions of area dorsalis telencephali pars medialis, centralis, and dorsolateral complex; and completely removed area ventralis telencephali pars dorsalis, ventralis, and lateralis. In addition, the supracommissural nucleus and rNPP were lesioned specifically because both nuclei have been thought to be involved in courtship behavior and endocrine control of reproduction. This investigation demonstrated extensive fiber projections from telencephalic nuclei and/or rNPP to the hypothalamus. Lesioned telencephalon and/or rNPP projected bilaterally to nucleus preopticus and the suprachiasmatic nucleus and unilaterally to the following tuberal nuclei: nucleus anterior tuberis, and the lateral hypothalamic nucleus. A much larger fiber projection to the inferior lobe nuclei was also observed with a large contralateral as well as ipsilateral input.

Peptidergic innervation of caudal neurosecretory neurons.

Luteinizing hormone releasing hormone (LHRH)-like immunoreactivity was observed in neurosecretory neurons at mesencephalic levels of the poecilid brain. This nucleus of peptide producing neurons has been shown to project to the spinal cord neurosecretory complex in this species. LHRH-like immunoreactivity was localized in fibers and terminals surrounding caudal neurosecretory cells. This investigation suggests that there is a mesencephalic descending LHRH innervation of caudal neurosecretory neurons.

Cytology of brain stem neurons projecting to the caudal neurosecretory complex: an HRP-electron microscopic study.

Brain stem projections to the neurons in the caudal neurosecretory complex (CNC) of Poecilia sphenops (molly) have been studied with HRP retrograde tracing. Using light microscopic procedures, HRP-labelled neurons were located in the reticular nucleus of the medulla (RMN) and in the vicinity of the nucleus of the medial longitudinal fascicle (NMLF). The present study was undertaken to examine the cytology of the brain stem neurons that project to the caudal neurosecretory complex using combined HRP-electron microscopic methods. Cells in the midbrain NMLF containing HRP-filled profiles were located bilaterally close to the midline and just beneath the ependyma. HRP reaction product was found additionally in the neurosecretory cells of the midbrain dorsal tegmental magnocellular nucleus (DTMN) located dorsal to the NMLF. Cells containing HRP-labelled profiles were also seen in the RMN and in neurons dorsal-lateral to the RMN. This latter group of neurons contained small dense core vesicles in addition to HRP labelled dense bodies.

Telencephalic terminals in the major retinal synaptic lamina of the goldfish optic tectum.

Light and electron microscopic degeneration studies were used to examine the telencephalotectal pathway in goldfish. Both techniques showed that each telencephalic lobe sent bilateral projections to several tectal laminae. Degenerating synaptic terminals and fibers were observed in the major retinal projection lamina as well as in other tectal laminae. The terminals contained round to oval synaptic vesicles, asymmetric synapses and contacted relatively small postsynaptic profiles.

Retinal terminals in the goldfish optic tectum: identification and characterization.

Retinal terminal profiles in the goldfish optic tectum were identified electron microscopically after (1) labeling with horseradish peroxidase and (2) in the early stages of degeneration in short-term eye enucleates. All labeled terminals shared certain common morphological characteristics which were identical to those of a population of terminals in normal tecta. Terminals of this type disappeared 30 days after enucleation of the contralateral eye. Retinal terminal presynaptic profiles were characterized by (1) round and oval synaptic vesicles; (2) mitochondria with irregular, randomly oriented cristae, large intracristal spaces, dilated membrane spaces, and primarily light matrices; (3) a wide range in profile area, 0.06-6.82 micrometers2; (4) large numbers of synaptic vesicles per profile area (168 +/- 33 synaptic vesicles per micrometers2; (5) asymmetric synapses; and (6) multiple synaptic contacts (1.46 +/- 0.73 per terminal profile). The postsynaptic elements included both dendritic and, less commonly, pleomorphic vesicle-containing profiles. The majority of postsynaptic dendritic profiles were small (0.01-0.40 micrometers2). Serial synaptic contacts were occasionally seen. The combination of vesicular and mitochondrial morphology (1 and 2 above) was necessary and sufficient to establish the retinal origin of a terminal, but use of such criteria would underestimate the number of retinotectal terminals by omitting those which did not have a mitochondrion in the plane of section. The number of such terminals was calculated from independent measurements, and the total number of retinal terminal profiles per area of neuropil was estimated.

Caudal neurosecretory system synaptic morphology following deafferentation: an electron microscopic degeneration study.

The caudal neurosecretory system of Poecilia sphenops (molly) is an isolated population of neurosecretory cells located in the caudal most aspect of the teleost spinal cord. The structure of this neuroendocrine system is favorable for studies on the synaptic control of neurosecretory mechanisms. Little is known about the detailed synaptology of the system. Morphological and electrophysiological reports have shown that the caudal neurosecretory system is linked to higher brain centers by descending spinal projections. To examine the synaptology of the descending synaptic input, surgical deafferentation was performed by microsuction removal of a segment of spinal cord rostral to the caudal system. The degeneration of axon terminals was studied at various times following deafferentation and compared to control synaptology. Based on vesicle content and morphology, three axon terminal types were found in the caudal neurosecretory system. These terminals formed axosomatic, axodendritic, and axoaxonic synaptic contacts. Following deafferentation, axon terminals with dense-cored vesicles and boutons with round clear vesicles degenerated as evidenced by the electron dense dark reaction and the electron lucent reaction respectively. This suggested that at least two different types of axon terminals arise from the descending projection to the caudal neurosecretory system and that two different neurotransmitters may be influencing the neurosecretory activity of these cells.

Supraependymal cells in the third ventricle of Poecilia sphenops.

The ependymal surface of the third ventricle in the hypothalamus of the black molly (Poecilia sphenops) was examined by scanning electron microscopy. Supraependymal cells were observed and could be categorized generally into two varieties, Types 1 and 2. P. sphenops is a representative of the teleostean family Poeciliidae which has been utilized as an effective neuroendocrine model. The presence of supraependymal cells in the hypothalamus ventricular space must be considered in future studies on hypothalamic-pituitary interaction.

Brain stem innervation of the caudal neurosecretory system.

The innervation of the caudal neurosecretory system of Poecilia sphenops (black molly) was studied by use of the retrograde horseradish peroxidase (HRP) method. The structure of the caudal neurosecretory system in this species was well suited for application of HRP procedures. Acrylamide/HRP gel implants were placed in the nucleus of the caudal neurosecretory system. Two neuronal groups which contained HRP filled cells were found in the brains tem. Bilateral projections originate from the dorsal tegmentum of the midbrain and the reticular nucleus of the medulla.

Ependymal cells of the filum terminale in fish (Poecilia sphenops) adapted to freshwater and saltwater: electron microscopic study.

The ependymal lining of the central canal of the filum terminale and spinal cord in the vicinity of the caudal neurosecretory system in P. sphenops was examined in this study. Two general cell types based on shape and location were observed in the ependymal lining: cuboidal ependyma located in dorsal aspects of the filum terminal and columnar to pseudostratified ependymal cells found in ventrolateral and ventral aspects of the filum terminale. Comparison of the ependymal lining was made in animals adapted to saltwater and freshwater. In animals adapted to saltwater there was an increase in the basal infolding of the cell membrane of the dorsal cuboidal ependyma. Infolding of the basal cell membrane is a phenomenon shared by cells known to participate in transport of electrolytes. Since a possible functional relationship between the ependyma of the third ventricle and median eminence has been shown, in future studies on the osmoregulatory function of the caudal neurosecretory system the ependymal lining of the central canal in this region should be considered.

The caudal neurosecretory system of Poecilia sphenops (Poeciliidae).

The caudal neurosecretory system of the molly, Poecilia sphenops (Poeciliidae) was studied by light and electron microscopy. In this species the cell bodies form a focal nuclear group in the caudal spinal cord. The neurosecretory cells are in contact with glial elements, axon terminals, and the lumen of the central canal. The axons of the neurosecretory cells form a definitive tract, which leaves the spinal cord proper to penetrate a well defined neurohemal organ, the urophysis. The urophysis contains an abundance of neurosecretory granules within the neurosecretory axonal processes. This study is the first ultrastructural study of the caudal neurosecretory system in this family of fishes, which has been used as a neuroendocrine model. This species acclimates easily to the laboratory aquarium and may be most suitable for further studies on the effects of changes in external salinity on the caudal neurosecretory system.