Light and electron microscopic immunocytochemical analysis of the noradrenaline innervation of the rat visual cortex

Summary Immunocytochemistry with an antiserum against noradrenaline was used to examine the organization and morphology of noradrenergic axons in the rat visual cortex. Observations with the light microscope confirmed earlier reports concerning the distribution pattern of noradrenergic fibres, and provided some further clues about their intracortical organization. Particularly striking was the finding of fibres which followed an oscillating course within the boundaries of layers II–IV as they ran in the mediolateral direction. Examination of the morphological characteristics of noradrenaline-containing axon terminals in serial ultrathin sections has provided further evidence that the vast majority (87.6%) form conventional synapses in the visual and frontoparietal cortex, and has given clues about the postsynaptic elements involved in these synaptic contacts; they are, in decreasing frequency, spines, dendritic shafts of various diameters, and pyramidal and non-pyramidal somata. In addition, a few labelled terminals were visualized in close association with intracerebral capillaries.     

Light and electron microscopic immunocytochemical analysis of the serotonin innervation of the rat visual cortex

Summary The serotonin afferents of the rat visual cortex were examined immunocytochemically at the light and electron microscopic levels. Immunoreactive fibres were typically thin, tortuous and varicose. Occasionally, some thicker fibres were found. The orientation of labelled axons varied according to laminar position, with fibres running parallel to the pial surface present mainly in layers I and VI, and radially oriented fibres prominent in layers II and III. Branches arising from horizontal or radially oriented fibres were seen to form irregularly shaped loops particularly in layers IV and V. The density of innervation and the prevailing axonal orientation in each cortical layer were similar in both coronal and parasagittal planes.The ultrastructural features of serotonin-labelled axon terminals were examined in single and serial ultrathin sections. While in single sections the majority did not exhibit synaptic specializations, extensive serial section analysis showed that virtually all of these terminals were engaged in junctional complexes. Postsynaptic elements were spines and dendritic shafts, including pyramidal cell apical dendrites, with both symmetrical and asymmetrical membrane specializations. In axospinous synapses, the labelled terminals were usually adjacent to unstained axon terminals contacting the same postsynaptic element.     

Somatostatin immunoreactive neurons in rat visual cortex: A light and electron microscopic study

Summary Somatostatin immunoreactive neurons in rat visual cortex were examined in the light and electron microscopes using an antibody to the tetradecapeptide form of somatostatin. Somatostatin immunoreactive neurons were found to belong only to non-pyramidal classes. They are of five main types: multipolar neurons with either thin or thick dendrites; small and large bipolar neurons; bitufted neurons; horizontal neurons; and neurons in the subcortical white matter. Of the immunoreactive neurons, multipolar neurons are the most common and account for 30% of the population, while bipolar and bitufted neurons make up 25% and 15% of the immunoreactive population, respectively; the least common somatostatin immunoreactive neurons are the horizontal and subcortical white matter neurons. Occasional multipolar neurons with thick dendrites have a prominent ascending dendrite so that they resemble pyramidal cells in the light microscope, but electron microscopic examination confirms that, like all other somatostatin-positive cells, they are non-pyramidal neurons, for they have both symmetric and asymmetric synapses on their cell bodies.Somatostatin-positive neurons are distributed among all the cortical layers and the subcortical white matter but they are more common in two laminae, one coinciding with layer II/III and the other with layers V and VI. The multipolar and bipolar neurons are distributed in similar proportions in these upper and lower cortical laminae, while bitufted neurons are more common in upper laminae and horizontal neurons are predominantly located in layer VI.     

Light and electron microscopic immunocytochemical analysis of the dopamine innervation of the rat visual cortex

Summary The dopaminergic innervation of the rat primary (area 17) and secondary (areas 18 and 18a) visual cortical areas was examined immunocytochemically using an antiserum directed against dopamine. This innervation was characterized by the differential density of the respective afferents within individual visual areas. Area 18, especially its rostral part, was observed to receive a considerable amount of dopaminergic axons, whereas areas 17 and 18a were sparsely innervated. The innervation of all layers of area 18 seemed to consist to a considerable extent of axonal branches of radial fibres ascending from layer VI to layer I. At the ultrastructural level, dopamine profiles were found to display similar characteristics in all visual areas. Dopamine labelled axon-terminals and axonal varicosities, examined in single and serial ultrathin sections, were seen to form primarily asymmetrical synaptic contacts with dendritic profiles. These observations suggest a specific innervation of cytoarchitectonically distinct cortical regions by dopaminergic axons.     

A quantitative electron microscopic study of atypical structures in normal human cerebral cortex.

Biopsy samples of the cerebral cortex from four normal human brains were examined in the electron microscope for the presence of abnormalities related to neurones and neuroglia. Atypical forms of axons, axon terminals and dendrites, many of them similar to those described in a variety of pathological and experimental material, were found to occur in small but appreciable numbers. Neuroglia exhibiting atypical inclusions occurred but were much less common. Physiologically altered neuronal perikarya were not encountered apart from one neuronal death. An area of 3.6 x 10(5) mu2 was scanned from each brain and atypical structures were categorised and counted. The possible implications of the presence of these structures in normal brain in discussed and the need for neuropathologists and neuroanatomists to be aware of the existence of atypical forms of neuronal processes in normal human and animal brains is emphasised.     

Synaptic connections of enkephalin-immunoreactive nerve terminals in the neostriatum: a correlated light and electron microscopic study

Summary Two different antisera to leucine-enkephalin were used to study the localization of enkephalin-like immunoreactive material in the neostriatum and globus pallidus of the rat, by means of the unlabelled antibody-enzyme method. Thin immunoreactive varicose fibres are scattered throughout the neostriatum. In the ventral striatum, fibres come together and follow a relatively straight course for several micrometers, forming tube-like structures which can be traced to cell bodies; these cell bodies are completely surrounded by immunoreactive fibres. Occasional immunoreactive varicose fibres are also found close to another type of neuron throughout the whole neostriatum.Examination by electron microscopy of immunoreactive structures that had been identified first in the light microscope, showed that each of the nearly 200 varicosities examined was a vesicle-containing bouton that formed a synaptic contact. Rarely were asymmetrical synaptic contacts found between immunoreactive boutons and dendritic spines. All other synapses formed by enkephalin-immunoreactive boutons were symmetrical. Two types of postsynaptic neuron were identified; the first type was a medium-sized neuron with the ultrastructural features of a typical striatal spiny neuron. The second type had a larger perikaryon surrounded by numerous immunoreactive varicosities that were found to be boutons forming symmetrical synapses. The long dendrites of this second type of neuron likewise received a dense input of immunoreactive boutons forming symmetrical synapses; such ensheathed dendrites were found to be the tube-like structures seen in the light microscope. The ultrastructural features of these neurons, notably a highly indented nucleus, were those of a rare type of striatonigral neuron. In the globus pallidus, all the enkaphalin-immunoreactive boutons studied formed symmetrical synapses with ensheathed dendrites and perikarya that were similar to the latter type of postsynaptic neuron in the neostriatum. Axo-axonic synapses involving immunoreactive boutons were not seen in our material.The results are consistent with the view that enkephalin-like substances may be synaptic transmitters in the neostriatum and that they may have different actions according to the nature of the postsynaptic target. The finding that one type of neostriatal neuron, and a very similar neuron in the globus pallidus, receives multiple enkephalin-immunoreactive boutons all over its perikaryon and along its dendrites indicates a potentially important role of enkephalin in the convergence of information within the neostriatum and pallidum on to output neurons.     

Heterogeneous actions of serotonin on interneurons in rat visual cortex

The effects of serotonin (5-HT) on excitability of two cortical interneuronal subtypes, fast-spiking (FS) and low threshold spike (LTS) cells, and on spontaneous inhibitory postsynaptic currents (sIPSCs) in layer V pyramidal cells were studied in rat visual cortical slices using whole-cell recording techniques. Twenty-two of 28 FS and 26 of 35 LTS interneurons responded to local application of 5-HT. In the group of responsive neurons, 5-HT elicited an inward current in 50% of FS cells and 15% of LTS cells, an outward current was evoked in 41% of FS cells and 81% of LTS cells, and an inward current followed by an outward current in 9% of FS cells and 4% LTS cells. The inward and outward currents were blocked by a 5-HT(3) receptor antagonist, tropisetron, and a 5-HT(1A) receptor antagonist, NAN-190, respectively. The 5-HT-induced inward and outward currents were both associated with an increase in membrane conductance. The estimated reversal potential was more positive than -40 mV for the inward current and close to the calculated K(+) equilibrium potential for the outward current. The 5-HT application caused an increase, a decrease, or an increase followed by a decrease in the frequency of sIPSCs in pyramidal cells. The 5-HT(3) receptor agonist 1-(m-chlorophenyl) biguanide increased the frequency of larger and fast-rising sIPSCs, whereas the 5-HT(1A) receptor agonist (+/-)8-hydroxydipropylaminotetralin hydrobromide elicited opposite effects and decreased the frequency of large events. These data indicate that serotonergic activation imposes complex actions on cortical inhibitory networks, which may lead to changes in cortical information processing.     

A transmission electron microscopic study of microglia/macrophages in the hippocampal cortex and neocortex following chronic exposure to valproate

In chronic administration of sodium valproate to rats, significant disorders of structural integrity of the hippocampal gyrus and the neocortex of the temporal lobe, observed in the last two stages of the experiment (after 9 and 12 months), coexisted with increased number of microglial cells and, especially after 12 months, with intense phagocytic activity within these cells. At the ultrastructural level, phagocyte microglial cells were hypertrophied with several broadened processes. Their cytoplasm contained rich lysosomal apparatus, numerous lipofuscin-like structures, lipid droplets and multilaminated bodies. The nuclei of these cells were characteristic oval or round and sometimes triangle in shape with dense and highly clumped heterochromatin, distinctly accumulated under nuclear envelope, and sparse euchromatin. Microglia/macrophages were frequently present in a close vicinity of changed neuronal somata and also close to the altered elements of the neuropil pyramidal layer of the cortex. Microglial response may, together with abnormalities in neurones, astroglia and blood-brain barrier, play a significant role in the development of experimental valproate encephalopathy.     

Synaptic relationships between a multipolar stellate cell and a pyramidal neuron in the rat visual cortex. A combined Golgi-electron microscope study

Summary Two synapsing and impregnated neurons in the rat visual cortex have been examined by a combined Golgi-electron microscope technique in which the Golgi precipitate is replaced by gold particles. One of the neurons is a stellate cell with smooth dendrites and a well impregnated axon, while the other is a layer III pyramidal neuron. Light microscopy showed some boutons from the axonal plexus of the stellate cell closely apposed to the soma and dendrites of the pyramid and it was predicted that synapses were present at these sites. An electron microscopic examination of serial thin sections, in which the profiles of the impregnated neurons are marked by their content of gold particles, showed most of these predicted synapses to exist. Indeed, axon terminals of the stellate cell formed five symmetric synapses with the cell body of the pyramid, one with the apical dendritic shaft and three with basal dendrites. Reasons are given for believing these synapses to be inhibitory.In addition, it was found that one of the axon terminals of the stellate cell synapsed with one of that cell's own dendrites. The significance of this finding is discussed.     

Intrahypothalamic connections: An electron microscopic study in the rat

Summary Terminal degeneration within the hypothalamus was studied by electron microscopy 1 or 2 days (1) after carefully placed microlesions in the arcuate, anterior periventricular, ventromedial, premammillary and posterior hypothalamic nuclei and (2) after microlesions placed in the hypothalamus deafferented 3 weeks earlier.In the median eminence terminal degeneration was found after each of these lesions. Projections from the ventromedial nucleus reach the arcuate, suprachiasmatic, and anterior periventricular nuclei.Projections from the arcuate nucleus terminate in the medial preoptic, anterior periventricular, and ventromedial nuclei.After lesioning the premammillary nuclei degeneration was found in the supraoptic, arcuate, anterior hypothalamic and ventromedial nuclei.     

Age-related changes in capillaries of rat oral mucosa. A quantitative electron microscopic study

Ultrastructural age changes in capillaries of the buccal mucosa were examined in montages of cross sections made from electron micrographs at 37,500 x. Six rats aged 6 months and 6 aged 30 months were perfused with glutaraldehyde and conventional thin sections obtained. Two capillaries located within a connective tissue papilla were studied from each rat. Capillaries of the old group differed from those of the young group by statistically significant increases in several parameters. The endothelial cell was increased in thickness, especially in the vicinity of junctions. The frequency of pinocytotic vesicles/unit length of the cell circumference was nearly doubled. Junctions were of nearly double length and took a more oblique course. All parts of the basement membranes were thickened, though perhaps less than seems true in skin. In striking contrast to epidermis, the epithelium of oral mucosa undergoes no appreciable thinning with age. We suggest that the observed age increases in frequency of pinocytotic vesicles and in the length of junctions may facilitate blood/tissue exchange, thus compensating for impaired exchange due to the thickened basement membranes. These compensatory changes in conjunction with the unchanged size of the mucosal capillary bed in the aged rat (demonstrated previously) could explain the unchanged thickness of the oral epithelium.     

Lipofuscin in the cerebellar cortex of albino rats: An electron microscopic study

Summary The ultrastructure of autofluorescent, PAS-positive lipofuscin in Purkinje, granule, Golgi epithelial, basket and stellate, microglial and perivascular cells in the cerebellar cortex of senescent rats is described. The membrane-bounded pigment is composed of three elements: 1) electron-lucent homogeneous droplets, 2) a granular matrix and 3) intensely osmiophilic patches. The proportions of these three components vary between cell types and one can grossly differentiate a neuronal and a glial lipofuscin. The lipofuscin granules of stellate and perivscular cells are different from lipofuscin of other cerebellar neurons and glia. It can be concluded from these morphological observations that each cerebellar cell type has its distinct lipofuscin.Supported by the Deutsche Forschungsgemeinschaft La 184/5I would like to thank Mrs. v. Bronewski and Mr. H. Boffin for their technical assistance     

Developmental regulation of intracellular calcium by N-methyl-D-aspartate and noradrenaline in rat visual cortex.

The effects of N-methyl-D-aspartate and noradrenaline on intracellular Ca2+ concentration in slices of rat visual cortex were studied using a fluorescent indicator, Fura-2. Bath application of N-methyl-D-aspartate (1-100 microM) increased intracellular Ca2+ concentration in a dose-dependent manner, especially in layers II/III. Noradrenaline (1-100 microM) also increased intracellular Ca2+ concentration in a dose-dependent manner, especially in layers I and IV. However, the maximum increase in intracellular Ca2+ concentration after 100 microM noradrenaline application was less than half of that after 100 microM N-methyl-D-aspartate application in slices obtained from animals in the sensitive period. The effect of noradrenaline was most prominent in slices of the sensitive period, whereas the N-methyl-D-aspartate-induced intracellular Ca2+ concentration response decreased with age. Additive effects from application of both N-methyl-D-aspartate and noradrenaline on intracellular Ca2+ concentration were found only in the neonatal stage. Pharmacological experiments showed that alpha1-adrenergic receptors play a major role in the noradrenaline-induced intracellular Ca2+ concentration response, although both alpha2- and beta-adrenergic receptors were also partially involved. The release of Ca2+ from intracellular storage underlay the early phase of the noradrenaline-induced intracellular Ca2+ concentration response, while extracellular Ca2+ influxes contributed to the sustained phase. Experiments using a gliotoxin, fluorocitric acid, suggested that the function of glial cells is involved in the noradrenaline-induced increase of intracellular Ca2+ concentration. The larger intracellular Ca2+ concentration response to noradrenaline during the sensitive period may modulate the increase in intracellular Ca2+ concentration by N-methyl-D-aspartate to maintain a higher level of cortical plasticity during this period.