Gracile nucleus of streptozotocin-induced diabetic rats

Summary This study reports ultrastructural changes in the gracile nucleus of male Wistar rats after streptozotocin-induced diabetes. During the acute phase (3–7 days) degenerating electron-dense dendrites and axon terminals were dispersed in the neuropil. Degenerating dendrites were characterized by an electron-dense cytoplasm, swollen mitochondria, dilated endoplasmic reticulum and scattered ribosomes. Degenerating axon terminals were characterized by an electron-dense cytoplasm and clustering of small spherical agranular vesicles. Degenerating axon terminals may form part of a synaptic glomerulus with a central electron-dense dendrite, or they may form the central element of a synaptic glomerulus. These degenerating profiles were absent in the gracile nucleus of the 3 and 7 days insulin-treated post-streptozotocin rats. Macrophages were present in the neuropil and were in the process of engulfing neuronal elements. During the medium phase (1–6 months), most of the degenerating dendrites and axon terminals had been engulfed or removed by macrophages. During the late phase (9–12 months) a second wave of degeneration occurred in the gracile nucleus, similar to the acute phase. During the medium and late phases, dystrophic axonal profiles were also significantly increased in the rats after streptozotocin treatment.It is concluded that the ultrastructural changes observed in the gracile nucleus in the present study were the result of streptozotocin-induced diabetes rather than a toxic effect of streptozotocin, even in the acute phase.     

Quantitative immunogold analysis reveals high glutamate levels in retinal and cortical synaptic terminals in the lateral geniculate nucleus of the macaque

An immunogold procedure has been used on ultrathin sections of the parvo- and magnocellular layers of the dorsal lateral geniculate of the rhesus monkey to estimate quantitatively at the electron microscopic level the intensity of immunoreactivity to an antibody against glutamate over profiles of retinal, cortical, GABAergic synaptic terminals and glial cells. GABAergic terminals were identified directly by immunogold reactivity to a GABA antibody or by ultrastructural features. The results showed that in both of the main subdivisions of the geniculate the densities of immunogold particles over cortical and retinal terminals were about two- to three-fold higher than those over GABAergic terminals or glial profiles. In addition, cortical and retinal terminals showed higher positive correlations of glutamate immunogold particle densities to synaptic vesicle densities than did GABAergic terminals. These differences suggest higher and lower concentrations of glutamate corresponding to transmitter and metabolic pools of this amino acid in axon terminals of retinal and cortical origins versus GABAergic terminals, respectively, in the dorsal lateral geniculate nucleus of the macaque.     

Glutamatergic components of the retrosplenial granular cortex in the rat

The ultrastructural characteristics, distribution and synaptic relationships of identified, glutamate-enriched thalamocortical axon terminals and cell bodies in the retrosplenial granular cortex of adult rats is described and compared with GABA-containing terminals and cell bodies, using postembedding immunogold immunohistochemistry and transmission electron microscopy in animals with injections of cholera toxin- horseradish peroxidase (CT-HRP) into the anterior thalamic nuclei. Anterogradely labelled terminals, identified by semi-crystalline deposits of HRP reaction product, were approximately 1 microm in diameter, contained round, clear synaptic vesicles, and established asymmetric (Gray type I) synaptic contacts with dendritic spines and small dendrites, some containing HRP reaction product, identifying them as dendrites of corticothalamic projection neurons. The highest densities of immunogold particles following glutamate immunostaining were found over such axon terminals and over similar axon terminals devoid of HRP reaction product. In serial sections immunoreacted for GABA, these axon terminals were unlabelled, whereas other axon terminals, establishing symmetric (Gray type II) synapses were heavily labelled. Cell bodies of putative pyramidal neurons, containing retrograde HRP label, were numerous in layers V-VI; some were also present in layers I-III. Most were overlain by high densities of gold particles in glutamate but not in GABA immunoreacted sections. These findings provide evidence that the terminals of projection neurons make synaptic contact with dendrites and dendritic spines in the ipsilateral retrosplenial granular cortex and that their targets include the dendrites of presumptive glutamatergic corticothalamic projection neurons.     

Ultrastructural correlates of synapse withdrawal at axotomized neuromuscular junctions in mutant and transgenic mice expressing the Wld gene

We carried out an ultrastructural analysis of axotomized synaptic terminals in Wld(s) and Ube4b/Nmnat (Wld) transgenic mice, in which severed distal axons are protected from Wallerian degeneration. Previous studies have suggested that axotomy in juvenile (< 2 months) Wld mice induced a progressive nerve terminal withdrawal from motor endplates. In this study we confirm that axotomy-induced terminal withdrawal occurs in the absence of all major ultrastructural characteristics of Wallerian degeneration. Pre- and post-synaptic membranes showed no signs of disruption or fragmentation, synaptic vesicle densities remained at pre-axotomy levels, the numbers of synaptic vesicles clustered towards presynaptic active zones did not diminish, and mitochondria retained their membranes and cristae. However, motor nerve terminal ultrastructure was measurably different following axotomy in Wld transgenic 4836 line mice, which strongly express Wld protein: axotomized presynaptic terminals were retained, but many were significantly depleted of synaptic vesicles. These findings suggest that the Wld gene interacts with the mechanisms regulating transmitter release and vesicle recycling.     

Pericapillary and distant axon terminals in the nuclei of the cat amygdala: a morphometric study

According to some ultrastructural studies, the pericapillary axon terminals in the central nervous system (CNS) are functionally connected with the capillary vessel wall. Thus, it may be expected that the population of pericapillary axon terminals will be morphologically distinct from the terminals at a further distance from the capillary walls. To test this hypothesis, morphometrical analysis of 3,048 axon terminals was performed, comparing terminals situated in the close vicinity of the capillary vessel with those at a distance from the vessels in the lateral, basal, medial, central and cortical nuclei of the amygdaloid body of eight cats. The cross-sectional area and circumference of each identified axon terminal profile were measured, and the shape of synaptic vesicles and the presence of synaptic contacts and granular vesicles were recorded. The statistical evaluation of results was performed by means of the Newman-Keuls' test, Wilcoxon's test, Fisher's contingency table test and the test for two coefficients of structure. The morphometric examination revealed two ultrastructurally distinct groups of axon terminals, pericapillary and distant terminals, in all the nuclei of the amygdaloid body. The differentiating features were the shape of the synaptic vesicles, the number of synaptic contacts, and the size of the axon terminals. These results further support the hypothesis of a functional connection between axon terminals and the capillary vessel wall in the CNS.     

Different kinds of axon terminals forming symmetric synapses with the cell bodies and initial axon segments of layer II/III pyramidal cells. II. Synaptic junctions

Summary Four different types of axon terminals form symmetric synapses with the cell bodies and initial axon segments of pyramidal cells in layer II/III of rat visual cortex. One type belongs to chandelier cells, and the other three kinds of terminals have origins that have not been established yet. These latter are referred to as large, medium-sized and dense terminals. The purpose of the present study was to examine the synaptic junctions formed by all four types of terminal. The synapses formed by the chandelier cell terminals are readily recognized in thin sections because of the characteristic features of both the terminals and the initial axon segments, which are the neuronal elements postsynaptic to them. In en face views of these axo-axonal synapses the junctions can be seen to have presynaptic dense projections that form a grid in which they are triagonally spaced, and have an average centre-to-centre spacing of 84 nm. As an ensemble the projections form the presynaptic grid, which usually has an oval or round outline, but may be notched on one side where projections are absent. The synaptic junctions of the large, medium-sized and dense terminals were examined by making reconstructions of the terminals from serial thin sections. It was found that at the interfaces between the axon terminals and the cell bodies of pyramidal cells, several separate synaptic junctions may be present, in addition to a number of puncta adhaerentia. Thus, there may be as many as five separate synaptic junctions and as few as one. It was also found that while the proportion of the area of the synaptic interface occupied by synaptic junctions was between 12% and 26% for dense terminals, for medium sized terminals it was 10–15%, and for the one large terminal reconstructed it was only 8%. Thus, there can be multiple synaptic junctions between each of these types of axon terminals and a pyramidal cell, and because many of the terminals forming symmetric junctions are boutons en passant, a number of vesicle release sites exist between the presynaptic axon and its postsynaptic partner. The axon terminals forming symmetric synapses in the cerebral cortex are assumed to be inhibitory, and consequently it is suggested that this arrangement of multiple release sites is designed to ensure that stimulation of the presynaptic axon results in an effective level of hyperpolarization of the postsynaptic neuron.     

Glutamatergic components of the retrosplenial granular cortex in the rat

The ultrastructural characteristics, distribution and synaptic relationships of identified, glutamate-enriched thalamocortical axon terminals and cell bodies in the retrosplenial granular cortex of adult rats is described and compared with GABA-containing terminals and cell bodies, using postembedding immunogold immunohistochemistry and transmission electron microscopy in animals with injections of cholera toxin- horseradish peroxidase (CT-HRP) into the anterior thalamic nuclei. Anterogradely labelled terminals, identified by semi-crystalline deposits of HRP reaction product, were approximately 1 m in diameter, contained round, clear synaptic vesicles, and established asymmetric (Gray type I) synaptic contacts with dendritic spines and small dendrites, some containing HRP reaction product, identifying them as dendrites of corticothalamic projection neurons. The highest densities of immunogold particles following glutamate immunostaining were found over such axon terminals and over similar axon terminals devoid of HRP reaction product. In serial sections immunoreacted for GABA, these axon terminals were unlabelled, whereas other axon terminals, establishing symmetric (Gray type II) synapses were heavily labelled. Cell bodies of putative pyramidal neurons, containing retrograde HRP label, were numerous in layers V–VI; some were also present in layers I–III. Most were overlain by high densities of gold particles in glutamate but not in GABA immunoreacted sections. These findings provide evidence that the terminals of projection neurons make synaptic contact with dendrites and dendritic spines in the ipsilateral retrosplenial granular cortex and that their targets include the dendrites of presumptive glutamatergic corticothalamic projection neurons.     

Axon hillocks and initial segments in spinal trigeminal nucleus with emphasis on synapses including axo-axo-axonic contacts

Summary As a part of a continuing study of the feline spinal trigeminal nucleus, the fine structure and synaptic arrangements on the axon hillock and axon initial segment of neurons in this region are described here. Transmission electron microscopy has been used to characterize qualitatively the axon hillock and initial segment and associated synapses in pars interpolaris. Axon hillocks and initial segments are easily identified in continuity with somata or as isolated profiles in the neuropil, and they receive synaptic contacts: these we regard as axo-axonic. The presynaptic terminals contain either mainly round or mainly flattened synaptic vesicles and have Type I (asymmetric) or Type II (symmetric) thickenings respectively at their contacts with the axon hillock or initial segment. I report here also the unusual arrangement of three separate axons in a serial synaptic complex. Some of the round vesicle Type I contacts onto the axon hillock-initial segment region also receive Type II contacts from one or more flattened vesicle terminals, thus formingan axo-axo-axonic complex. These flattened vesicle terminals lack the usual features of a presynaptic dendrite. It has been shown that in this nucleus some round vesicle terminals, especially those postsynaptic to flattened vesicle terminals, are primary afferents from the periphery. Therefore the round vesicle terminal presynaptic to the axon hillock-initial segment region, some of which are included in the axo-axo-axonic complex may also be a primary afferent directly contacting the spike generator area of the relay neuron and under presynaptic control of a flattened vesicle synapse. The latter may possibly be an intrinsic contact. This strategic situation of round vesicle terminals and the axo-axo-axonic complex at the axon hillock or initial segment has major implications relevant to the overall output of these neurons.     

Effect of light deprivation upon the morphology of axon terminals in the dorsal lateral geniculate nucleus of mouse: an electron microscopical study using serial sections

Two populations of morphologically different large axon terminals have been observed electron microscopically in the dorsal lateral geniculate nucleus of mice raised in complete darkness from birth up to 19 days of age. One population includes larger terminals indistinguishable from the large terminals present in control animals, i.e. they have round synaptic vesicles, rather pale mitochondria, membrane saculae, coated vesicles, and asymmetric contacts with encrusted dendritic spines of portions of dendrites of geniculo-cortical relay neurons. The other population includes large terminals which also have asymmetric contacts with encrusted dendritic spines or portions of dendrites of geniculo-cortical relay neurons, but they show darker mitochondria, absence of both membrane saculae and coated vesicles, and significantly higher synaptic vesicle density and smaller size than the large control ones. We suggest that the latter population of terminals could be inactive due to the absence of visual input.     

包埋前免疫电镜双标技术在神经解剖学研究中的应用

目的　在超微结构水平观察两种神经递质在纤维终末内的共存或一种神经递质与其相应受体之间的关系。　方法　包埋前免疫电镜双重标记技术———酶标法和免疫金 银标记法相结合的方法。　结果　在免疫反应双重标记的纹状体切片上 ,电镜下观察到大量的SP样 (过氧化物酶免疫反应产物 )阳性终末和SP受体 (SPR ,免疫金 银标记颗粒 )样阳性神经元的胞体和树突 ,同时可见部分SP样阳性轴突终末分别与SPR样阳性神经元的胞体或树突形成对称性轴 体或轴 树突触联系。而在三叉神经脊束核尾侧亚核切片上 ,电镜下可观察到大量的两种囊泡膜谷氨酸转运体 ,即DNPI样 (过氧化物酶免疫反应产物 )和VGluT1样 (免疫金 银标记颗粒 )阳性轴突终末 ,同时还观察到DNPI样和VGluT1样双标的轴突终末与阴性树突形成非对称性突触。　结论　包埋前免疫电镜双重标记技术敏感性较高 ,组织的抗原性保存好 ,特别是在神经解剖学研究中 ,用于研究两种神经递质在同一个细胞或终末内的共存或分析神经递质与其相应受体之间的联系中有独到之处。     

Synaptic organization of the cat entopeduncular nucleus with special reference to the relationship between the afferents to entopedunculothalamic projection neurons: An electron microscope study by a combined degeneration and horseradish peroxidase tracing technique

The synaptic organization of the feline entopeduncular nucleus was studied electron microscopically. After horseradish peroxidase injections into the ventral anterior and ventral lateral nuclear complex of the thalamus, normal axon terminals synapsing with entopedunculothalamic projection neurons were classified into four types on the basis of the size and shape of synaptic vesicles in them, and types of the postsynaptic membrane differentiation. Type I and type II axon terminals were characterized by symmetrical synaptic contacts, and large ovoid or small ovoid synaptic vesicles, respectively. Type II axon terminals were further classified into two subtypes as to their sizes: one was small (IIa), the other large (IIb). Type III and type IV axon terminals were characterized by asymmetrical synaptic contacts, and large ovoid or small ovoid synaptic vesicles, respectively.

To determine the origin of each type of terminal, electrolytic lesions of the caudate nucleus or the subthalamic nuclear region were combined with horseradish peroxidase injections into the thalamus or the subthalamic nuclear region. After electrolytic lesions of the caudate nucleus, degeneration was seen in type I axon terminals contacting entopedunculothalamic projection neurons. Following electrolysis or horseradish peroxidase injection into the subthalamic nuclear region, type IIa and type IV axon terminals showed degenerations or horseradish peroxidase labelling. Such terminals also synapsed with entopedunculothalamic projection neurons. It was demonstrated that these projection neurons relay the striatal or subthalamic inputs directly to the thalamus. After horseradish peroxidase injection into the thalamus, many labelled type II axon terminals were observed to synapse with entopedunculothalamic projection neurons. Type III axon terminals were left unchanged throughout these experiments. In addition, the entopeduncular neuron was observed to receive convergent inputs from both the caudate nucleus and probably the subthalamic nucleus. Axoaxonal synapses were also found to be involved in the synaptic triad.

These results indicate that type I axon terminals originate from the caudate nucleus, part of type IIa and type IV axon terminals originate from the subthalamic nucleus or caudal to the subthalamic nuclear region, and part of type IIa and type IIb terminals come from intrinsic axon collaterals.     

Synaptic vesicle pools at diaphragm neuromuscular junctions vary with motoneuron soma, not axon terminal, inactivity

Both spinal hemisection (SH) at C2 and tetrodotoxin (TTX) phrenic nerve blockade result in diaphragm muscle paralysis and inactivity of the phrenic axon terminals. However, phrenic motoneuron somata are inactive with SH but remain active with TTX phrenic nerve blockade. Neuromuscular transmission failure with repeated activation decreases following SH and increases following TTX phrenic nerve blockade, suggesting that matching (or mismatching) of somal and synaptic inactivities of phrenic motoneurons differentially regulates synaptic vesicle pools at diaphragm neuromuscular junctions. At individual type-identified rat diaphragm presynaptic terminals, the size of the releasable pool of synaptic vesicles was analyzed by fluorescence confocal microscopy of N-(3-triethylammoniumpropyl)-4-(6-(4-(diethylamino)phenyl)hexatrienyl) pyridinium dibromide (FM4-64) uptake and synaptic vesicle density at active zones was determined using transmission electron microscopy. After 14 days of SH and TTX-induced diaphragm muscle inactivity, neuromuscular junction size was not different at type I or IIa fibers, but increased at type IIx and/or IIb fibers (by 51% in SH and 35% in TTX) compared with control. With SH, synaptic vesicle pool size and density increased at presynaptic terminals innervating type I or IIa fibers (17 and 63%, respectively; P<0.001) and type IIx and/or IIb fibers (41 and 31%, respectively; P<0.001) when compared with controls. Following TTX, synaptic vesicle pool size and density decreased by 64 and 17%, respectively, at presynaptic terminals innervating type I or IIa fibers, and by 50 and 36%, respectively, at type IIx and/or IIb fibers (P<0.001, for all comparisons). Thus, matching motoneuron soma and axon terminal inactivity (SH) increases the size and density of releasable synaptic vesicle pools at adult rat diaphragm neuromuscular junctions. Mismatching motoneuron soma and axon terminal inactivities (TTX) results in converse presynaptic adaptations. Inactivity-induced neuromuscular plasticity reflects specific adaptations in the size and density of synaptic vesicle pools that depend on motoneuron soma rather than axon terminal (or muscle fiber) inactivity.     

Ultrastructure of synaptic connections of a bimodal pacemaker giant neuron in the central nervous system of Helix pomatia L.

Following intracellular labelling with horseradish peroxidase, the arborization and synaptic connections of the bimodal pacemaker giant neuron (RPal) of Helix pomatia were investigated in the right parietal and visceral ganglia. The RPal neuron possesses extensive axonal branching, the elements of which could be observed and traced within the entire neuropil region of both ganglia. The main axonal branches showed further arborization. The thin axon processes enter the synaptic neuropil, where they receive numerous synapses. At least six ultra-structurally different terminals form synaptic contacts on peroxidase-labelled axon processes of the cell. On the basis of their vesicle and granule content, they are likely to contain different neurotransmitters. Some intraganglionic efferent contacts of the RPal neuron were also observed.It is suggested that, besides its peripheral efferent connections, this cell might also serve as an interneuron.     

Inhibition of acetylcholinesterase accelerates axon terminal withdrawal at the developing rat neuromuscular junction

Summary In developing skeletal muscles, the rate at which superfluous innervation is lost from the endplates depends on the general level of neuromuscular activity. Whether it is activity of the presynaptic or postsynaptic structures (or both) that is critical is not well established. In this work, we transitorily inhibited the AChE of soleus muscle in postnatal rats, in order to increase postsynaptic activity, without directly altering activity of the nerve terminals. We then followed the time course of disappearance of axon terminals from the endplates of treated and normal muscles, using electron-microscope techniques. Three hours after inhibition of AChE, the muscle fibres exhibited local supercontracture and ultrastructural damage in the region of the endplate, consistent with local elevation of Ca²⁺ levels. At the same time, small electron-opaque vesicles, apparently of muscular origin, appeared in the synaptic cleft. The nerve terminals, however, were entirely normal in number and appearance. One day after treatment, endplates of esteraseinhibited muscles showed accelerated loss of nerve terminals, compared to endplates of normally developing muscles. No further loss of nerve terminals occurred, once AChE activity returned at the endplate. These results suggest that the rate at which superfluous nerve terminals retract from the developing neuromuscular junction is regulated by the level of activation of the muscle. It seems likely that activity of postsynaptic sites may similarly regulate changes in innervation patterns, in other developing or adapting neuro-neuronal or neuro-effector systems.     

Injury-induced remodelling and regeneration of the ribbon presynaptic terminalin vitro

Summary The neuronal response to axonal injury may relate to the type of insult incurred. Recently, neuritic and presynaptic varicosity regeneration by isolated adult salamander photoreceptors was demonstrated. We have used this system to compare the rod photoreceptor response to two types of injury:denervation/detargeting, the removal of pre-and postsynaptic partners from the axon terminal, andaxotomy, the removal of the axon terminal itself. Cells were followed with time-lapse video microscopy for 24–48h in culture and immunolabelled for SV2 or synaptophysin to identify synaptic vesicle-containing varicosities. Although all injured cells responded with regenerative growth, denervated/detargeted photoreceptors (i.e. neurons which retain their axon terminal) grew 80% more processes and fourfold more presynaptic varicosities than axotomized neurons. In cells which retained their original axon and terminal, varicosity formation generally began with axon retraction. Retraction was followed by elaboration of a lamellipodium and, by 48 h, development of varicosity-bearing neurites from the lamellipodium. Synaptic vesicle protein localization in denervated/detargeted cells paralleled axon terminal reorganization. Axotomized cells, in contrast, lacked synaptic vesicle protein immunoreactivity during this period. To detect synaptic protein synthesis, photoreceptors were examined for colocalization of synaptic vesicle protein with rab6, a Golgi marker, by confocal microscopy. As expected, synaptic vesicle protein staining was present in the Golgi complex during regeneration; however, in cells with an axon, new synaptic vesicle protein-labelled varicosities were found at early stages, prior to the appearance of immunolabel in the Golgi complex. The data demonstrate remarkable plasticity in the ribbon synapse, and suggest that in adult rod cells with an intact axon terminal, synaptic vesicle protein synthesis is not a prerequisite for the formation of new presynaptic-like terminals. We propose that preexisting axonal components are reutilized to expedite presynaptic renewal as an early response to denervation/detargeting.     

Vesicle recycling at ribbon synapses in the finely branched axon terminals of mouse retinal bipolar neurons

In retinal bipolar neurons, synaptic ribbons mark the presence of exocytotic active zones in the synaptic terminal. It is unknown, however, where compensatory vesicle retrieval is localized in this cell type and by what mechanism(s) excess membrane is recaptured. To determine whether endocytosis is localized or diffuse in mouse bipolar neurons, we imaged FM4-64 to track vesicles in cells whose synaptic ribbons were tagged with a fluorescent peptide. In synaptic terminals, vesicle retrieval occurred at discrete sites that were spatially consistent over multiple stimuli, indicative of endocytotic “hot spots.” Retrieval sites were spatially correlated with fluorescently labeled synaptic ribbons. Electron microscopy (EM) analysis of bipolar cell terminals after photoconversion of internalized FM dye revealed that almost all of the dye was contained within vesicles ∼30 nm in diameter. Clathrin-coated vesicles were observed budding from the plasma membrane and within the cytosol, and application of dynasore, a dynamin inhibitor, arrested membrane retrieval just after the budding stage. We conclude that synaptic vesicles in the fine branches of mouse bipolar axon terminals are retrieved locally near active zones, at least in part via a clathrin-mediated pathway.     

Suprachiasmatic nucleus neurons immunoreactive for vasoactive intestinal polypeptide have synaptic contacts with axons immunoreactive for neuropeptide Y: an immunoelectron microscopic study in the rat

An electron microscopic study showed by using a dual immunolabeling technique that in the suprachiasmatic nucleus of the rat, axon terminals immunoreactive for neuropeptide Y (NPY) made synaptic contacts upon neurons immunoreactive for vasoactive intestinal polypeptide (VIP). Diaminobenzidine (DAB)-labeled NPY axon terminals made synaptic contacts on silver-gold-labeled VIP perikarya and dendritic processes. The presynaptic NPY terminals contained many small clear vesicles and a few cored vesicles labeled with DAB chromogen. At the synaptic portion, a symmetrical thickening of the pre- and post-synaptic membranes was evident.     

Synaptic junctions between sympathetic axon terminals and pinealocytes in the monkey Macaca fascicularis

Summary The distribution of axon terminals in the pineal gland of monkeys was studied by electron microscopy. Numerous terminals bearing small pleomorphic agranular and dense-cored vesicles were localized in the perivascular space and among the pinealocytes in the parenchyma in normal monkeys. Following bilateral superior cervical ganglionectomy, they underwent degenerative changes, including the accumulation of glycogen masses, appearance of dense residual bodies and the displacement of synaptic vesicles. Some of these degenerating terminals showed synaptic contacts with the cell bodies of pinealocytes. At the synaptic junction the postsynaptic membrane was thickened asymmetrically. Examples of synaptic contacts were most frequently observed in 5 and 7 days postoperative animals. In the longer surviving (30 days) monkey, most of the axon terminals showed round agranular vesicles, and they were mainly presynaptic to the intrapineal ganglion cells with some of the pinealocytes. They remained structurally unchanged following the resection of both the superior cervical ganglia. A few axon terminals containing small dense-cored vesicles appeared to have survived the initial insult, but some of their vesicles appeared swollen 30 days after the operation. It is concluded from this study that some of the pinealocytes are under the influence by the postganglionic neurons in the superior cervical ganglia through direct synaptic contacts. The intrapineal ganglion cells are postsynaptic to fibres originating exclusively from the central nervous system. Some of these fibres, however, may be presynaptic directly to pinealocytes.     

Ultrastructural analysis of the central terminals of primary sensory neurones labelled by transganglionic transport of bandeiraea simplicifolia I-isolectin B4

In this study the ultrastructural appearance of primary sensory neurones labelled by the injection of the plant lectin Bandeiraea simplicifolia I-isolectin B(4) (BSI-B(4)) into a peripheral nerve has been examined in the rat. Electron microscopy of the somata of retrogradely labelled neurones showed the lectin to be associated with the inner surface of cytoplasmic vesicles, supporting the premise that the uptake of BSI-B(4) into sensory neurones is by the process of receptor-mediated endocytosis. Light and electron microscopic analysis of the spinal cord revealed transganglionically transported lectin in unmyelinated axons in the dorsolateral funiculus and axon terminals concentrated mainly within lamina II of the dorsal horn. Detailed analysis of 1377 of these axon terminals revealed that the majority were glomerular in shape and surrounded by up to 14 other unlabelled profiles. These findings suggest that primary sensory neurones which transganglionically transport BSI-B(4) have a synaptic ultrastructure similar to that which has been previously reported for unmyelinated primary sensory neurones. Moreover, it appears that the axon terminals of these neurones are subjected to extensive modulation. Examination of the vesicle content of lectin labelled axon terminals revealed that the majority contained small agranular vesicles while large granular vesicles were observed only occasionally. These findings support the suggestion that the populations of neurones expressing binding sites for BSI-B(4) are fairly distinct from those containing neuroactive peptides. In conclusion, the results of the current study suggest that the lectin BSI-B(4) can be used as a histological marker for a subpopulation of small diameter primary sensory neurones and provide further evidence for the potential of this lectin as a useful tool in the study of pain.     

Parvalbumin-immunoreactive neurons in the entorhinal cortex of the rat: localization, morphology, connectivity and ultrastructure

Summary We studied the distribution, morphology, ultrastructure and connectivity of parvalbumin-immunoreactive neurons in the entorhinal cortex of the rat. Immunoreactive cell bodies were found in all layers of the entorhinal cortex except layer I. The highest numbers were observed in layers II and III of the dorsal division of the lateral entorhinal area whereas the lowest numbers occurred in the ventral division of the lateral entorhinal area, Most such neurons displayed multipolar configurations with smooth dendrites. We distinguished a type with long dendrites and a type with short dendrites. We also observed pyramidal immunoreactive neurons. A dense plexus of immunoreactive dendrites and axons was prominent in layers II and III of the dorsal division of the lateral entorhinal area and the medial entorhinal area. None of the parvalbuminimmunoreactive cells became retrogradely labelled after injection of horseradish peroxidase into the hippocampal formation. By electron microscopy, immunoreactivity was observed in cell bodies, dendrites, myelinated and unmyelinated axons and axon terminals. Immunoreactive dendrites and axons occurred in all cortical layers. We noted many myelinated immunoreactive axons. Immunoreactive axon terminals were medium sized, contained pleomorphic synaptic vesicles, and established symmetrical synapses. Both horseradish peroxidase labelled and unlabelled immunonegative cell bodies often received synapses from immunopositive axon terminals arranged in baskets. Synapses between immunoreactive axon terminals and unlabelled dendritic shafts and spines were abundant. Synapses with initial axon segments occurred less frequently. In addition, synaptic contacts were present between immunopositive axon terminals and cell bodies and dendrites. Thus, the several types of parvalbumin-containing neuron in the entorhinal cortex are interneurons, connected to one another and to immunonegative neurons through a network of synaptic contacts. Immunonegative cells projecting to the hippocampal formation receive axo-somatic basket synapses from immunopositive terminals. This connectivity may form the morphological substrate underlying the reported strong inhibition of cells in layers II and III of the entorhinal cortex projecting to the hippocampal formation.