Catecholaminergic innervation of pyramidal and GABAergic nonpyramidal neurons in the rat hippocampus. Double label immunostaining with antibodies against tyrosine hydroxylase and glutamate decarboxylase

This study describes the catecholaminergic innervation of rat hippocampal neurons at the electron microscopic level by using an antibody against tyrosine hydroxylase (TH) and immunocytochemical techniques. In a first series of experiments, the course and distribution as well as the synaptic contacts of TH-immunoreactive fibers were analyzed with the peroxidase-antiperoxidase (PAP) method. Next, peroxidase immunostaining of TH fibers was combined with glutamate decarboxylase (GAD) immunostaining, using avidinated ferritin as a second electrondense marker. Our results demonstrate that TH-immunostained terminals establish asymmetric synaptic contacts with spines of pyramidal neurons, and symmetric synaptic contacts with cell bodies and dendritic shafts of ferritin-labeled GAD-immunoreactive nonpyramidal cells.     

GABA-immunohistochemistry as a label for identifying types of local interneurons and their synaptic contacts in the antennal lobes of the American cockroach

Summary Synaptic contacts between GABA-immunoreactive neurons, antennal receptor fibers and non-GABA-immunoreactive neurons in the glomerular neuropil of the antennal lobes have been identified by means of a combination of (i) immunohistochemical labeling and (ii) labeling of afferent fibers of the antenna by experimentally induced degeneration. Characteristic contacts of these neurons are: a) Serially arranged polysynaptic contacts between degenerated antennal fibers, GABA-immunoreactive neurons and non-GABA-immunoreactive neurons. b) Monosynaptic contacts between degenerated antennal fibers and non-GABA-immunoreactive neurons. c) Reciprocal synaptic contacts between immunostained and non-stained neurons and synaptic contacts between individual GABA-immunoreactive neurons. d) Synaptic output contacts of GABA-immunoreactive neurons with degenerated antennal fibers.GABA-immunoreactive neuron profiles in the glomeruli are assigned to multiglomerular local interneurons (Distler 1989a); non-immunolabeled profiles may be assigned to projection neurons and other not yet identified interneurons.     

GABA-immunohistochemistry as a label for identifying types of local interneurons and their synaptic contacts in the antennal lobes of the American cockroach

Synaptic contacts between GABA-immunoreactive neurons, antennal receptor fibers and non-GABA-immunoreactive neurons in the glomerular neuropil of the antennal lobes have been identified by means of a combination of (i) immunohistochemical labeling and (ii) labeling of afferent fibers of the antenna by experimentally induced degeneration. Characteristic contacts of these neurons are: a) Serially arranged polysynaptic contacts between degenerated antennal fibers, GABA-immunoreactive neurons and non-GABA-immunoreactive neurons. b) Monosynaptic contacts between degenerated antennal fibers and non-GABA-immunoreactive neurons. c) Reciprocal synaptic contacts between immunostained and non-stained neurons and synaptic contacts between individual GABA-immunoreactive neurons. d) Synaptic output contacts of GABA-immunoreactive neurons with degenerated antennal fibers. GABA-immunoreactive neuron profiles in the glomeruli are assigned to multiglomerular local interneurons (Distler 1989a); non-immunolabeled profiles may be assigned to projection neurons and other not yet identified interneurons.     

Immunocytochemical observations of corticotropin-releasing-factor-containing neurons in the rat hypothalamus with special reference to neuronal communication

Synapses between neurons with corticotropin-releasing-factor-(CRF)-like immunoreactivities and other immunonegative neurons in the hypothalamus of colchicine-treated rats, especially in the paraventricular nucleus (PVN) and the supraoptic nucleus (SON) were observed by immunocytochemistry using CRF antiserum. The immunoreactive nerve cell bodies and fibers were numerous in both the PVN and the SON. The CRF-containing neurons had synaptic contacts with immunonegative axon terminals containing a large number of clear synaptic vesicles alone or combined with a few dense-cored vesicles. We also found CRF-like immunoreactive axon terminals making synaptic contacts with other immunonegative neuronal cell bodies and fibers. And since some postsynaptic immunonegative neurons contained many large neurosecretory granules, they are considered to be magnocellular neurosecretory cells. These findings suggest that CRF functions as a neurotransmitter and/or modulator in addition to its function as a hormone.     

The intrastriatal injection of thrombin in rat induced a retrograde apoptotic degeneration of nigral dopaminergic neurons through synaptic elimination

We have performed intrastriatal injection of thrombin and searched for distant effects in the cell body region. In striatum, thrombin produced a slight loss of striatal neurons as demonstrated by neural nuclei immunostaining – a non-specific neuronal marker – and the expression of glutamic acid decarboxylase 67 mRNA, a specific marker for striatal GABAergic interneurons, the most abundant phenotype in this brain area. Interestingly, striatal neuropil contained many boutons immunostained for synaptic vesicle protein 2 and synaptophysin which colocalize with tyrosine hydroxylase (TH), suggesting a degenerative process with pre-synaptic accumulation of synaptic vesicles. When we studied the effects on substantia nigra, we found the disappearance of dopaminergic neurons, shown by loss of TH immunoreactivity, loss of expression of TH and dopamine transporter mRNAs, and disappearance of FluoroGold-labelled nigral neurons. The degeneration of substantia nigra dopaminergic neurons was produced through up-regulation of cFos mRNA, apoptosis and accumulation of α-synuclein shown by colocalization experiments. Thrombin effects could be mediated by protease-activated receptor 4 activation, as protease-activated receptor 4-activating peptide mimicked thrombin effects. Our results point out the possible relationship between synapse elimination and retrograde degeneration in the nigral dopaminergic system.     

Dopamine Synthesis Precedes Dopamine Uptake in Embryonic Rat Mesencephalic Neurons

We have measured [3H]dopamine ([3H]DA) uptake and tyrosine hydroxylase-immunopositive immunostaining in cells acutely dissociated from the embryonic ventral mesencephalon (MSC). DA and its metabolites as well as catechol-O-methyltransferase (COMT) and monoamine oxidase (MAO) activities were determined in homogenates taken from the MSC and striatum (STR). In the embryonic ventral MSC measurable DA and tyrosine hydroxylase (TH) immunostaining were present as early as embryonic day (E) 12.5. At E14 the number of TH+ neurons was about 50% of the values at E18. In the MSC, DA concentration increased sharply at E16 and reached a plateau before birth that was 10-fold lower than adult values. In the STR, DA was first detected at E16, suggesting that DA fibers reach the STR at this embryonic stage. High-affinity DA uptake appeared in the MSC only at E16, concomitantly with the arrival of DA fibers in the STR, increased sharply between E16 and E18, and reached a plateau before birth. This uptake mechanism was not selective for catecholamine uptake inhibitors. Thus, DA synthesis in the MSC preceded the onset of high-affinity uptake mechanism, which could be correlated to the beginning of striatal DA innervation. Measurable MAO and COMT activities were detected as early as E13 (MSC) and E15 (STR), but not DA metabolites, which appeared later. We conclude that the high-affinity DA uptake mechanism in MSC DA neurons develops coincident with the arrival of DA fibers to the STR. The sharp increase of DA uptake between E16 and E18 is due only in part to an increase in the number of TH+ cells. These results support the hypothesis that in vivo the target STR neurons regulate the maturation of MSC DA cells.     

Synaptic Contacts of Neurons of Fascia Dentata Transplants with Nonspecific Targets in Neocortex of Recipients

We carried out an electron microscopy study of possible synaptic contacts of the neurons of intracortical transplants of the rat brain fascia dentata with targets in the recipient somatosensory cortex. The axons of fascia dentata granular cell and their synaptic terminals could be easily identified in the neocortex due to their distinct morphological features (mossy fibers), although the fascia dentate cells normally do not interact with the neocortex. Thin nonmyelenized mossy fibers were found in both an intermediate zone between the transplant and brain and in the adjacent brain. Their presynaptic buds, like in situ, had large size and formed characteristic terminal, intraterminal, and en passant multiple synaptic contacts and desmosome-like junctions. The aberrant nerve fibers used perykaryons, dendrites of varying diameter, and dendrite spikes of the somatosensory cortex pyramidal neurons as postsynaptic targets in the neocortex. In addition to vacant spaces that appeared in the brain as a result of transplantation, the ingrowing axons induced the formation of additional contact sites: deep invaginations of the plasmalemma of perykaryons, somatic spikes, terminal branchings of dendrites, and dendritic outgrowths of complex branched shape. These aberrant contacts were characterized by the presence of polyribosomes, endoplasmic reticulum cisternae, and mitochondria in the postsynaptic loci. Osmiophility and extension of desmosome-like junctions were also enhanced in such synapses. Thus, it was shown that mossy fibers ingrowing in the recipient neocortex were capable of forming cell-to-cell contacts with signs of functional synapses to atypical cell targets.     

Synaptic contacts of neurons of fascia dentata transplants with nonspecific targets in neocortex of recipients

We carried out an electron microscopy study of possible synaptic contacts of the neurons of intracortical transplants of the rat brain fascia dentata with targets in the recipient somatosensory cortex. The axons of fascia dentata granular cell and their synaptic terminals could be easily identified in the neocortex due to their distinct morphological features (mossy fibers), although the fascia dentate cells normally do not interact with the neocortex. Thin nonmyelenized mossy fibers were found in both an intermediate zone between the transplant and brain and in the adjacent brain. Their presynaptic buds, like in situ, had large size and formed characteristic terminal, intraterminal, and en passant multiple synaptic contacts and desmosome-like junctions. The aberrant nerve fibers used perykaryons, dendrites of varying diameter, and dendrite spikes of the somatosensory cortex pyramidal neurons as postsynaptic targets in the neocortex. In addition to vacant spaces that appeared in the brain as a result of transplantation, the ingrowing axons induced the formation of additional contact sites: deep invaginations of the plasmalemma of perykaryons, somatic spikes, terminal branchings of dendrites, and dendritic outgrowths of complex branched shape. These aberrant contacts were characterized by the presence of polyribosomes, endoplasmic reticulum cisternae, and mitochondria in the postsynaptic loci. Osmiophility and extension of desmosome-like junctions were also enhanced in such synapses. Thus, it was shown that mossy fibers ingrowing in the recipient neocortex were capable of forming cell-to-cell contacts with signs of functional synapses to atypical cell targets.     

Synaptic relationships between GABA-immunoreactive neurons and an identified uniglomerular projection neuron in the antennal lobe of Periplaneta americana: a double-labeling electron microscopic study

Two types of central neurons in the antennal lobe of the American cockroach Periplaneta americana were labeled with a combination of two specific markers. Their synaptic contacts were characterized and their distribution on the neurons examined. A uniglomerular pheromone-sensitive projection neuron with dendritic arbor in the male-specific macroglomerulus (attractant neuron) was characterized physiologically by intracellular recording and then filled with biocytin, which was converted to a marker for this individual neuron by a preembedding procedure. In a postembedding procedure local, multiglomerular interneurons were marked by immunogold labeling of GABA. Two kinds of synaptic contacts were found on the attractant neuron. (i) Input synapses from GABA-immunoreactive profiles. There were many of these, which (together with results of previous studies) suggests that local interneurons mediate polysynaptic transmission from antennal receptor fibers to the projection neuron. (ii) Output synapses onto GABA-immunoreactive profiles and onto non-identified neurons. These contacts indicate that signals generated by the projection neurons in a given glomerulus are passed back to multiglomerular interneurons and hence are also transmitted to other glomeruli.     

Central Connectivity of Transient Receptor Potential Melastatin 8-Expressing Axons in the Brain Stem and Spinal Dorsal Horn

Transient receptor potential melastatin 8 (TRPM8) ion channels mediate the detection of noxious and innocuous cold and are expressed by primary sensory neurons, but little is known about the processing of the TRPM8-mediated cold information within the trigeminal sensory nuclei (TSN) and the spinal dorsal horn (DH). To address this issue, we characterized TRPM8-positive (+) neurons in the trigeminal ganglion and investigated the distribution of TRPM8+ axons and terminals, and their synaptic organization in the TSN and in the DH using light and electron microscopic immunohistochemistry in transgenic mice expressing a genetically encoded axonal tracer in TRPM8+ neurons. TRPM8 was expressed in a fraction of small myelinated primary afferent fibers (23.7%) and unmyelinated fibers (76.3%), suggesting that TRPM8-mediated cold is conveyed via C and Aδ afferents. TRPM8+ axons were observed in all TSN, but at different densities in the dorsal and ventral areas of the rostral TSN, which dominantly receive sensory afferents from intra- and peri-oral structures and from the face, respectively. While synaptic boutons arising from Aδ and non-peptidergic C afferents usually receive many axoaxonic contacts and form complex synaptic arrangements, TRPM8+ boutons arising from afferents of the same classes of fibers showed a unique synaptic connectivity; simple synapses with one or two dendrites and sparse axoaxonic contacts. These findings suggest that TRPM8-mediated cold is conveyed via a specific subset of C and Aδ afferent neurons and is processed in a unique manner and differently in the TSN and DH.     

Neuronal interactions between neuropeptide Y (NPY) and catecholaminergic systems in the rat arcuate nucleus as shown by dual immunocytochemistry

Several recent studies have suggested interactions between catecholamine (CA) and neuropeptide Y (NPY) neuronal systems in the rat brain. In order to obtain morphological evidence for such CA/NPY interactions in the arcuate nucleus, we have used a double immunostaining procedure using an anti-tyrosine hydroxylase (TH) antiserum as a marker for catecholamine neurons and an anti-NPY antiserum. This double staining, where the first staining is silver-gold intensified, was detectable at both light and electron microscopic levels. In semi-thin sections, a substantial overlap and close proximity of TH-immunopositive neurons and NPY neuronal elements could be seen within the arcuate nucleus. At the electron microscopic level, direct appositions between TH- and NPY-immunoreactive structures could be detected. These appositions were of axosomatic, axodendritic or axoaxonic types without any synaptic membrane differentiation. Moreover, direct appositions between NPY-immunoreactive structures have also been observed. This morphological study showing appositions between TH and NPY neuronal systems suggest direct interactions between these two systems in the arcuate nucleus.     

Synaptic relationships between GABA-immunoreactive neurons and an identified uniglomerular projection neuron in the antennal lobe of Periplaneta americana: a double-labeling electron microscopic study

Summary Two types of central neurons in the antennal lobe of the American cockroach Periplaneta americana were labeled with a combination of two specific markers. Their synaptic contacts were characterized and their distribution on the neurons examined. A uniglomerular pheromone-sensitive projection neuron with dendritic arbor in the male-specific macroglomerulus (attractant neuron) was characterized physiologically by intracellular recording and then filled with biocytin, which was converted to a marker for this individual neuron by a preembedding procedure. In a postembedding procedure local, multiglomerular interneurons were marked by immunogold labeling of GABA. Two kinds of synaptic contacts were found on the attractant neuron. (i) Input synapses from GABA-immunoreactive profiles. There were many of these, which (together with results of previous studies) suggests that local interneurons mediate polysynaptic transmission from antennal receptor fibers to the projection neuron. (ii) Output synapses onto GABA-immunoreactive profiles and onto non-identified neurons. These contacts indicate that signals generated by the projection neurons in a given glomerulus are passed back to multiglomerular interneurons and hence are also transmitted to other glomeruli.     

Intrinsic choroidal neurons in the duck eye receive sympathetic input: anatomical evidence for adrenergic modulation of nitrergic functions in the choroid

Intrinsic choroidal neurons (ICN) in the duck eye form an intramural ganglionic plexus that may subserve complex integrative functions. A key feature of such ganglia is an innervation by sympathetic postganglionic neurons. The present study was thus aimed at determining the sympathetic postganglionic innervation of ICN. Choroids were processed for double immunofluorescence labelling with the following markers: tyrosine-hydroxylase (TH)/nitric oxide synthase (nNOS), TH/galanin (GAL), dopamine-beta-hydroxylase (DBH)/vasoactive intestinal polypeptide (VIP), TH/DBH and DBH/alpha-smooth-muscle actin (alphaSMA), and for triple immunofluorescence labelling with VIP/DBH/TH. Epifluorescence and confocal laser scanning microscopy were used for evaluation. Immunoperoxidase staining for TH or DBH in combination with NADPH-diaphorase histochemistry was applied for electron microscopy. ICN spread over the entire choroid but were concentrated in an equatorial zone passing obliquely from naso-cranial to temporocaudal. More than 80% of nNOS-positive ICN showed close appositions of TH/DBH-immunoreactive varicose nerve fibres at the light-microscopic level, as could be confirmed by confocal laser scanning microscopy. Ultrastructurally, these appositions could be defined as both synapses or close contacts without synaptic specialisation. Vascular and non-vascular smooth muscle fibres also received TH/DBH-immunopositive innervation. Our findings suggest that most ICN receive a sympathetic input that might modulate their nitrergic effects upon vascular and non-vascular smooth muscle fibres in the choroid and that they may have more complex functions than merely being a simple parasympathetic relay.     

NPY and TH innervation in human choroidal whole-mounts

To determine the distribution of NPY and TH human choroidal innervation, choroidal whole-mounts were processed for indirect immunofluorescence. An antibody to a component of the neuronal cytoskeleton, neurofilament 200 kDa (NF-200) was used to identify neurons and axons. A double immunostaining was performed, antibodies against NF-200 being combined with antibodies against neuropeptide Y (NPY) and tyroxine hydroxylase (TH). Fibers containing both NPY and TH were distributed in three plexuses, one in the suprachoroid large-sized vessel layer, and two in the medium-sized vessel layer. Intrinsic choroidal neurons (ICNs) containing NPY and TH were observed in the suprachoroid. The TH(+) ICNs were located in the medium-sized vessel layer. Overall, NPY(+) and TH(+) ICNs were more frequent in the central temporal area, both in isolation and forming microganglia. We also detected small spindle elements intensely immunoreactive to TH(+) and distributed mainly in the suprachoroid from the equator to the periphery. In conclusion, the human choroid contains abundant NPY and TH nerve fibers related to chroroidal vascular structures; it further possesses NPY(+) and TH(+) ICNs which contribute to the choroidal self-regulation persisting after sympathetic denervation. Additionally, these ICNs may at least partially explain why the choroidal blood flow does not respond to the factors that influence systemic vascular control. The preferential location of these cells in the submacular area suggests that dysfunction or degeneration of these cells may be a factor in vascular pathologies found in ocular disease, such as diabetic macular edema or age-related macular degeneration.     

Neuronal and synaptic organization of the lateral geniculate nucleus of the tree shrew,Tupaia glis

Summary The ultrastructural study of the lateral geniculate nucleus (LGN) of the tree shrew (Tupaia glis) revealed two types of neurons: (1) a large thalamocortical relay cell (TCR), which may bear cilia, and (2) a small Golgi type-II interneuron (IN) with an invaginated nucleus. The narrow rim of pale cytoplasm of the IN contains fewer lysosomes and fewer Nissl bodies than the cytoplasm of the TCR. The IN perikarya, which in some cases establish somatosomatic contacts, frequently contain flattened or pleomorphic synaptic vesicles. The ratio of TCR to IN is 31.Three types of axon terminals were observed in the LGN. Two of them contain round synaptic vesicles but differ in size. The large RL boutons undergo dark degeneration after enucleation; they are the terminals of retino-geniculate fibers. The smaller RS boutons show dark degeneration after ablation of the visual cortex; they are the terminals of the cortico-geniculate fibers. The third type of bouton (F₁ does not degenerate after either intervention. The boutons of this type are filled with flattened vesicles and are believed to be intrageniculate terminals. F₂-profiles were interpreted as presynaptic dendrites of the IN. The characteristic synaptic glomeruli found in the LGN contain in their center an optic terminal. These optic terminals establish synaptic contacts with dendrites or spine-like dendritic protrusions of TCRs as well as with presynaptic dendrites. Synaptic triads were also seen. The distribution of the individual types of synaptic contacts in layers 3 and 4 was determined. Layer 4 contains only one third of the retino-geniculate synapses and of the synaptic contacts of F₁-terminals.     

An ultrastructural study of peripheral neurons and associated non-neural structures in the bivalve mollusc, Spisula solidissima

The ultrastructural morphology of peripheral neurons and associated structures in the bivalve mollusc, Spisula solidissima have been studied in an effort to describe the synaptic topography and to provide anatomical correlates of previous physiological observations. The somata of the peripheral neurons are located within the perineurium at branch points of the siphonal nerves. There are many fiber-fiber synaptic contacts which are characterized by isolated sites of contact with membrane specialization and unilateral accumulation of synaptic vesicles. There are also synaptic contacts involving the somata, both axo-somatic and somato-axonic, the two being distinguishable on the basis of the polarity of vesicle accumulation. All of the observed synaptic profiles were similar in appearance regardless of the neuron loci involved. Much of the non-synaptic soma surface is covered with processes of glial cells. Likewise, in many cases, individual fibers and groups of fibers are encased with glial processes. Unique clusters of membrane bound, pigment containing glial like cells occur throughout the nervous system of Spisula. The heterogeneous appearance of the inclusions and the presence of lysosome-like bodies in the cytoplasm of these cells suggest a possible phagocytic role.     

Specific distribution pattern of nerve fibers containing catecholamine-synthesizing enzymes, neuropeptide Y (NPY) and C-terminal flanking peptide of NPY (CPON) in the pineal gland of the chinchilla (Chinchilla laniger)--an immunohistochemical study

The sympathetic nerve fibers originating from the superior cervical ganglia and supplying the pineal gland play the most important role in the control of the pineal activity in mammals. NPY and CPON are also present in the majority of the pinealopetal sympathetic neurons. In this study, immunohistochemical techniques were used to demonstrate the existence and coexistence of tyrosine hydroxylase (TH), dopamine beta-hydroxylase (DbetaH) as well as NPY and CPON in the nerve fibers supplying the chinchilla pineal gland. Ten two-year-old female chinchillas housed in natural light conditions were used in the study. The pineals were fixed by perfusion. ABC immunohistochemical technique and immunofluorescence labelling method were employed. TH-immunoreactive (TH-IR) varicose nerve fibers were observed in the pineal gland as well as in the posterior commissural area. Within the chinchilla pineal gland, TH-IR nerve fibers were located in the capsule and connective tissue septa. Numerous varicose TH-IR branches penetrated into the parenchyma and formed a network showing the highest density in the proximal region of the gland. In the central and distal parts of the pineal parenchyma, a subtle network, composed of thin varicose nerve branches, was observed. Double immunostaining revealed that the majority of TH-IR nerve fibers was positive for DbetaH or NPY. TH- and DbetaH-positive neuron-like cells were observed in the proximal region of the gland. The pattern of pineal innervation immunoreactive to CPON was similar to the innervation containing NPY, TH and DbetaH. The chinchilla intrapineal innervation containing TH, DbetaH, NPY and CPON is characterized by the higher density in the proximal part of the gland than in the middle and distal ones. The specific feature of the chinchilla pineal is also the presence of single TH/DbetaH-immunoreactive neuron-like cells in the proximal part of the gland.     

Light- and electron-microscopic study of synaptic connections in the paracervical ganglion of the female rat: special reference to calcitonin gene-related peptide-, galanin- and tachykinin (substance P and neurokinin A)-immunoreactive nerve fibers and terminals

Nerve fibers and varicosities in the pelvic paracervical ganglia (PG) are immunoreactive for the neuropeptides calcitonin gene-related peptide, galanin, and the tachykinins substance P and neurokinin A. Many of these fibers and varicosities are capsaicin-sensitive, originate in dorsal root ganglia and, thus, are considered to be primary afferent fibers. Numerous immunoreactive varicosities are pericellular to principal neurons in the PG. The present study examines the ultrastructure of calcitonin gene-related peptide-, galanin-, substance P-, and neurokinin A-immunoreactive nerve fibers and varicosities in the ganglia to determine their relationships to principal neurons and their synaptic connectivity. Paracervical ganglia of female rats were processed for light-microscopic immunohistochemistry using antisera against synapsin I, as a nerve terminal marker, and microtubule-associated protein-2 to define soma and dendrites. The rationale for performing this co-immunohistochemical analysis was to reveal the relationship between nerve endings and principal neurons. Synapsin I endings were predominantly axosomatic with fewer being axodendritic. Other ganglia were processed for electron-microscopic immunohistochemistry using both standard immunogold and peroxidase-anti-peroxidase procedures. Unmyelinated fibers and varicosities immunoreactive for calcitonin gene-related peptide, galanin, and the tachykinins were routinely observed in the interstitium between neuron somas. Numerous immunoreactive axon profiles were present in small groups that were ensheathed by Schwann cells. Immunoreactive fibers and varicosities were also observed within the satellite-cell sheath of the neuron soma and often intimately associated with the membrane of the soma, somal protrusions, or with the proximal part of a dendrite. Membrane specializations, indicative of synaptic contacts, between the fibers and the principal neurons were observed. It is suggested that these peptide-immunoreactive sensory fibers and varicosities are involved in regulation of activity in the PG.     

Fine structural analysis of calcitonin gene-related peptide in the mouse inferior olivary complex

Climbing fiber afferents to the cerebellum, from the inferior olivary complex, have a powerful excitatory effect on Purkinje cells. Changes in the responsiveness of olivary neurons to their afferent inputs, leading to changes in the firing rate or pattern of activation in climbing fibers, have a significant effect on the activation of cerebellar neurons and ultimately on cerebellar function. Several neuropeptides have been localized in both varicosities and cell bodies of the mouse inferior olivary complex, one of which, calcitonin gene related peptide (CGRP), has been shown to modulate the activity of olivary neurons. The purpose of the present study was to investigate the synaptic relationships of CGRP-containing components of the caudal medial accessory olive and the principal olive of adult mice, using immunohistochemistry and electron microscopy. The vast majority of immunoreactive profiles were dendrites and dendritic spines within and outside the glial boundaries of synaptic glomeruli (clusters). Both received synaptic inputs from non-CGRP labeled axon terminals. CGRP was also present within the somata of olivary neurons as well as in profiles that had cytological characteristics of axons, some of which were filled with synaptic vesicles. These swellings infrequently formed synaptic contacts. At the LM level, few, if any, CGRP-immunoreactive climbing fibers, were seen, suggesting that CGRP is compartmentalized within the somata and dendrites of olivary neurons and is not transported to their axon terminals. Thus, in addition to previously identified extrinsic sources of CGRP, the widespread distribution of CGRP within olivary somata and dendrites identifies an intrinsic source of the peptide suggesting the possibility of dendritic release and a subsequent autocrine or paracrine function for this peptide within olivary circuits.     

Colocalization of tyrosine hydroxylase and Fos in the male Syrian hamster brain following different states of arousal

In an investigation of the role that central tyrosine hydroxylase-(TH) containing neurons play in copulation in the male Syrian hamster, The induction of Fos protein was used as an index of neuronal activation. With a double immunoperoxidase technique, the activation of TH neurons was compared in hamsters from three experimental groups: (1) mated in a new cage; (2) handled controls placed in a new cage, and (3) unhandled controls. Although mating selectively induces Fos production in the medial amygdaloid nucleus (Me), more than half of the TH neurons in Me (a region outside of the classical catecholamine systems) expressed Fos equally in all of the experimental groups. In the paraventricular hypothalamic nucleus (PVN), TH neurons were activated equivalently in mated and handled control animals compared to unhandled controls. TH neurons in the neucleus of the solitary tract (NST) were also activated in handled control animals, and mating further enhanced the level of Fos immunostaining in these neurons above both groups of nonmated animals. Although not quantified, co-localization of Fos and TH was also observed in all experimental groups in the olfactory bulbs and the interfascicular nucleus, and in the horizontal limb of the diagonal band of Broca and the cerebral cortex, regions which contain TH neurons but are not part of the classically described TH cell groups. Few, if any, TH neurons in other catecholaminergic brain regions, such as the substantia nigra and locus coeruleus, produced Fos in any of the experimental groups. These results suggest that TH neurons in the PVN and NST may be activated during different states of arousal, and that nonclassical TH neurons in the amygdala produce high levels of Fos even in unstimulated animals. 1994 John Wiley & Sons, Inc.