Generation of resonance-dependent oscillation by mGluR-I activation switches single spiking to bursting in mesencephalic trigeminal sensory neurons.

The primary sensory neurons supplying muscle spindles of jaw-closing muscles are unique in that they have their somata in the mesencephalic trigeminal nucleus (MTN) in the brainstem, thereby receiving various synaptic inputs. MTN neurons display bursting upon activation of glutamatergic synaptic inputs while they faithfully relay respective impulses arising from peripheral sensory organs. The persistent sodium current (IN aP ) is reported to be responsible for both the generation of bursts and the relay of impulses. We addressed how IN aP is controlled either to trigger bursts or to relay respective impulses as single spikes in MTN neurons. Protein kinase C (PKC) activation enhanced IN aP only at low voltages. Spike generation was facilitated by PKC activation at membrane potentials more depolarized than the resting potential. By injection of a ramp current pulse, a burst of spikes was triggered from a depolarized membrane potential whereas its instantaneous spike frequency remained almost constant despite the ramp increases in the current intensity beyond the threshold. A puff application of glutamate preceding the ramp pulse lowered the threshold for evoking bursts by ramp pulses while chelerythrine abolished such effects of glutamate. Dihydroxyphenylglycine, an agonist of mGluR1/5, also caused similar effects, and increased both the frequency and impedance of membrane resonance. Immunohistochemistry revealed that glutamatergic synapses are made onto the stem axons, and that mGluR1/5 and Nav1.6 are co-localized in the stem axon. Taken together, glutamatergic synaptic inputs onto the stem axon may be able to switch the relaying to the bursting mode.

[THE ORGANIZATION OF PROJECTIONS OF MIDBRAIN LATERAL TEGMENTAL NUCLEI THE TO BRAIN BASAL GANGLIA IN DOGS].

The organization of the projections of midbrain lateral tegmental nuclei (peripeduncular nucleus, paralemniscal nucleus, nucleus of the brachium of inferior colliculus) to functionally diverse nuclei of the basal ganglia system was studied in dogs (n = 34) by the method of retrograde axonal transport of horse-radish peroxidase. It was found that the midbrain nuclei studied were involved in functionally different circuits, containing the basal ganglia as their components. These nuclei innervate the regions of the putamen, globus pallidus, cuneate nucleus, subcuneate nucleus, which are the motor or the limbic structures on the basis of their predominant connections with the motor or the limbic brain nuclei, and also regions of the caudate nucleus, nucleus accumbens, entopeduncular nucleus, compact part of the pedunculopontine nucleus, which receive the projections from the functionally various structures. The analysis of Nissl-stained frontal sections allowed to refine the anatomical topography of the individual nuclei of the midbrain lateral tegmentum. The cholinergic nature of their neurons was demonstrated based on of the positive histochemical reaction to NADPH diaphorase.

[The analysis of the possible pathways of carrying information and its integration in the projection systems, connecting the basal ganglia with the rostromedial tegmental nucleus dog's brain].

The structural foundation of processing of the information in the basal ganglia morphofunctional system was studied on the basis of the analysis of the projections between the separate parts of the rostral and thecaudal compartments of the rostromedial tegmental nucleus (RMTN) and the functionally different segments of the striatal and the pallidal structures, which were investigated by the method of the retrograde axonal transport. The elements of the topic, showing to opportunity of segregated carrying of the information between the limbic segments RMTN and the limbic striopallidal fields were revealed in the organization of the projections between the named structures in dog. But the convergence of the projection fibers, proceeding from the neurons of the functionally different parts of RMTN observed in the majority of striopallidal structures that testify about the opportunity of the integration of the functionally different information in them. The revealed labeled RMTN "reticular" neurons with sparsely branching dendrites and long axons, which are projected to the striopallidal structures, also testify about the integrative function of the investigated nucleus. The possible ways of carrying of the motor and the limbic information and its integration in the structures of the studied system and opportunity of the utilization of the received dates for making of the new models, which allow to understand better the mechanism of basal ganglia functioning in normal and pathological conditions are discussed.

[Organization of the projections of the substructures of deep mesencephalic nucleus to the striatum in the dog brain].

Method of retrograde axonal transport of horseradish peroxidase was used to study the organization of the projections of the individual substructures of the deep mesencephalic nucleus complex to the functionally diverse regions of striatal structures of the dog brain (n=20). It was shown that the projection fibers of the neurons of the nucleus profundus mesencephali were directed to the ventro-lateral segment of the nucleus caudatus and the ventral segment of the putamen. The dorsal segment of the putamen obtains the fibers from the neurons of the nucleus cuneiformis, while the lateral segment of the nucleus accumbens received them from the neurons of the nucleus subcuneiformis. The possible pathways for conduction of the functionally diverse information and its integration in the investigated projection systems are discussed.

Matrix metalloproteinase-12 deficiency ameliorates the clinical course and demyelination in Theiler's murine encephalomyelitis.

Matrix metalloproteinases (MMPs) are a family of extracellular proteases involved in the pathogenesis of demyelinating diseases like multiple sclerosis (MS). The aim of the present study was to investigate whether MMPs induce direct myelin degradation, leukocyte infiltration, disruption of the blood-brain barrier (BBB), and/or extracellular matrix remodeling in the pathogenesis of Theiler's murine encephalomyelitis (TME), a virus-induced model of MS. During the demyelinating phase of TME, the highest transcriptional upregulation was detected for Mmp12, followed by Mmp3. Mmp12 (-/-) mice showed reduced demyelination, macrophage infiltration, and motor deficits compared with wild-type- and Mmp3 knock-out mice. However, BBB remained unaltered, and the amount of extracellular matrix deposition was similar in knock-out mice and wild-type mice. Furthermore, stereotaxic injection of activated MMP-3, -9, and -12 into the caudal cerebellar peduncle of adult mice induced a focally extensive primary demyelination prior to infiltration of inflammatory cells, as well as a reduction in the number of oligodendrocytes and a leakage of BBB. All these results demonstrate that MMP-12 plays an essential role in the pathogenesis of TME, most likely due to its primary myelin- or oligodendrocyte-toxic potential and its role in macrophage extravasation, whereas there was no sign of BBB damage or alterations to extracellular matrix remodeling/deposition. Thus, interrupting the MMP-12 cascade may be a relevant therapeutic approach for preventing chronic progressive demyelination.

The inhibitory influence of the lateral habenula on midbrain dopamine cells: ultrastructural evidence for indirect mediation via the rostromedial mesopontine tegmental nucleus.

The lateral habenula (LHb) provides an important source of negative reinforcement signals to midbrain dopamine (DA) cells in the substantia nigra and ventral tegmental area (VTA). This profound and consistent inhibitory influence involves a disynaptic connection from glutamate neurons in the LHb to some population of γ-aminobutyric acid (GABA) cells that, in turn, innervates DA neurons. Previous studies demonstrated that the GABA cells intrinsic to the VTA receive insufficient synaptic input from the LHb to serve as the primary source of this intermediate connection. In this investigation, we sought ultrastructural evidence supporting the hypothesis that a newly identified region of the brainstem, the rostromedial mesopontine tegmental nucleus (RMTg), is a more likely candidate for inhibiting midbrain DA cells in response to LHb activation. Electron microscopic examination of rat brain sections containing dual immunoreactivity for an anterograde tracing agent and a phenotypic marker revealed that: 1) more than 55% of the synapses formed by LHb axons in the RMTg were onto GABA-labeled dendrites; 2) more than 80% of the synapses formed by RMTg axons in the VTA contacted dendrites immunoreactive for the DA synthetic enzyme tyrosine hydroxylase; and 3) nearly all RMTg axons formed symmetric synapses and contained postembedding immunoreactivity for GABA. These findings indicate that the newly identified RMTg region is an intermediate structure in a disynaptic pathway that connects the LHb to VTA DA neurons. The results have important implications for understanding mental disorders characterized by a dysregulation of reward circuitry involving LHb and DA cell populations.

The cholinergic mesopontine tegmentum is a relatively neglected nicotinic master modulator of the dopaminergic system: relevance to drugs of abuse and pathology.

The mammalian mesopontine tegmentum (MPT) contains two cholinergic nuclei, the pedunculopontine tegmental nucleus (PPTg) and the laterodorsal tegmental nucleus (LDTg). These provide the cholinergic innervation of, among other brain areas, the dopaminergic A9 and A10 cell groups. Their axons are thus the source of endogenous acetylcholine (ACh) acting on somato-dendritic acetylcholine receptors in the substantia nigra (SN) and ventral tegmental area (VTA). The anatomy, physiology, functional and pathological implications of these interactions with the nicotinic subtype of acetylcholine receptors (nAChRs) are discussed with a view of the important role of the MPT as a master regulator of nicotinic dopaminergic signalling in the brain, including for nicotine addiction.

Premotor circuits controlling eyelid movements in conjunction with vertical saccades in the cat: II. interstitial nucleus of Cajal.

Vertical saccadic eye movements are accompanied by concurrent eyelid movements in the same direction. The interstitial nucleus of Cajal (InC) controls eye position for vertical eye movements and may also control saccade-related lid position as well. This study investigates whether the InC serves as a premotor center for eyelid saccades, by employing dual-tracer methods in cats to label both the projections of the InC and the motoneurons supplying the levator palpebrae superioris (LPS) muscle, which lie in the caudal central subdivision (CCS) of the oculomotor complex. Injections of biotinylated dextran amine (BDA) into the InC anterogradely labeled axons that terminated bilaterally throughout the CCS and in the oculomotor nuclei proper. Labeled terminals lay in close association with labeled LPS motoneurons, which were retrogradely labeled following injections of wheat germ agglutinin-conjugated horseradish peroxidase (WGA-HRP) into the muscle. Ultrastructural investigation revealed that most terminals contained spherical vesicles and formed asymmetric synaptic contacts with the labeled motoneurons. These results strongly suggest that the InC monosynaptically controls lid movements in conjunction with vertical eye movements, including saccades. To identify the neurons of origin for this pathway, WGA-HRP injections were centered in the CCS. These experiments indicate that lid and eye motoneurons may share a common source of bilateral InC input. Thus, a common vertical position signal may be employed to maintain the lid and eye at appropriate elevations during fixation, such that the lid sits just above the pupil, allowing unobstructed vision, but at the ready to protect the cornea.

Laterodorsal tegmental projections to identified cell populations in the rat ventral tegmental area.

Projections from the laterodorsal tegmentum (LDT) to the ventral tegmental area (VTA) contribute to the activity of dopamine (DA) and GABA cells and, hence, to the affective and cognitive functions of this region. LDT afferents arise from neurochemically diverse cell types and mediate multiple functional influences. However, the VTA cell populations that receive LDT afferents are unknown and were investigated here by anterograde and retrograde tract-tracing in combination with immunocytochemistry to distinguish DA and GABA cells. Approximately 50% of the LDT to VTA pathway formed asymmetric, presumably excitatory synapses that innervated DA and GABA cells in rough proportion to their representation within the VTA. This portion of the LDT innervation appeared to selectively target DA but not GABA mesoaccumbens neurons and provide a relatively nonselective input to both DA and GABA mesoprefrontal cells. The remaining LDT axons formed symmetric, presumably inhibitory synapses with a different pattern of cellular targets that included a preferential input to GABA neurons of both mesoaccumbens and mesoprefrontal populations and an apparently selective innervation of mesoprefrontal and not mesoaccumbens DA neurons. These data suggest that the LDT mediates a convergent excitatory and inhibitory influence on both mesoprefrontal DA and GABA cells but a divergent impact on mesoaccumbens neurons that is likely to excite DA cells and inhibit GABA neurons. Combined with our previous description of prefrontal cortical afferents, our data also indicate that mesoaccumbens DA neurons receive putative excitatory drive from the LDT, whereas mesoprefrontal DA cells receive convergent excitation from both cortical and brainstem sources.

Synaptic organization of the rat parafascicular nucleus, with special reference to its afferents from the superior colliculus and the pedunculopontine tegmental nucleus.

The synaptic organization of afferents to the parafascicular nucleus (Pf) of the thalamus was studied in rats. In the Pf, three types of axon terminals were identified: the first type was a small terminal with round synaptic vesicles forming an asymmetric synapse, the second type was a large terminal with round synaptic vesicles forming an asymmetric synapse, and the third type was a terminal with pleomorphic vesicles forming a symmetric synapse. They were named SR, LR and P boutons, respectively. In order to determine the origin of these axon terminals, biotinylated dextran amine (BDA) was injected into the main afferent sources of the Pf, the superior colliculus (SC) and the pedunculopontine tegmental nucleus (PPN). Axon terminals from the SC were both SR and LR boutons which made synaptic contacts with somata and dendrites. PPN afferents were SR boutons, which made synaptic contacts with somata and smaller dendrites. Double-labeled electron microscopic studies, in which a retrograde tracer (wheat germ agglutinin conjugated to horseradish peroxidase: WGA-HRP) was injected into the striatum and an anterograde tracer (BDA) into the SC revealed that SC afferent terminals made synapses directly with Pf neurons that projected to the striatum. Another experiment was performed to find out whether two different afferents converged onto a single Pf neuron. To address this question, two different tracers were injected into the SC and PPN in a rat. Electron microscopically, both afferent terminals from the SC and PPN made synaptic contacts with the same dendrite. Our results prove that a single neuron of the rat Pf received convergent projections from two different sources.

Serotonergic innervation of the isthmo-optic nucleus of the pigeon centrifugal visual system. An immunocytochemical electron microscopic study.

The ultrastructural features of serotonergic fibers, terminals and synaptic contacts were studied with the pre-embedding immunocytochemical method in the isthmo-optic nucleus of the pigeon centrifugal visual system. The 5-HT immunoreactive (-ir) profiles were diffusely distributed and their density was low. The labeled axons were thin and unmyelinated (mean diameter=0.21+/-0.03 microm) though a few larger myelinated axons were observed (mean diameter=0.51+/-0.07 microm). The 5-HT-ir terminals or varicosities were small (diameter=0.71+/-0.54 microm) and contained small agranular synaptic vesicles (diameter=28.5+/-6.9 nm) and large granular vesicles (diameter=102.2+/-19.5 nm). The latter only constituted approximately 1% of the total profiles containing synaptic vesicles in the isthmo-optic nucleus. In single thin sections, only 5% of the 5-HT-ir varicosities exhibited an active asymmetrical zone synapsing upon dendritic profiles of centrifugal visual neurons. Calculations indicated that 17% of these 5-HT-ir varicosities were actually engaged in junctional synaptic relationships, whereas the remaining (83%) were nonjunctional. The data suggest that, within the isthmo-optic nucleus, 5-HT acts both at synaptic junctions (wiring transmission) and at a distance via the extracellular space (volume transmission). These 5-HT afferents could thus modulate the activity of the retinopetal neurons and visual information processing.

Distribution of terminals from pedunculopontine tegmental nucleus and synaptic organization in lateralis medialis-suprageniculate nucleus of cat's thalamus: anterograde tracing, immunohistochemical studies, and quantitative analysis.

The cat's lateralis medialis-suprageniculate nuclear complex (LM-Sg) in the thalamus receives input from various brain regions such as the superior colliculus, brain stem, and spinal cord, as well as from visual association cortex. In a previous study, we demonstrated that LM-Sg receives cholinergic fibers from the pedunculopontine tegmental nucleus (PPT) and that cholinergic terminals make synaptic contacts with the dendrites of glutamatergic projection neurons and of GABAergic interneurons (Hoshino et al., 1997). In this study, we investigate the distribution and the organization of PPT terminals by means of a combined anterograde tracer (biotinylated dextran amine, BDA) and immunohistochemical methods. When stained by acetylcholinesterase (AChE), the LM-Sg is not uniformly immunoreactive, but rather is patchily labeled and shows a streaming type of reactivity. The tissue content appears high in enzyme activity in AChE-positive zones and is much lighter in activity in AChE-negative zones. We compared the synaptic organization between AChE-positive and AChE-negative portions of the LM-Sg in separate groups of electron-microscopic material: four types of vesicle containing profiles (RS, RL, F1, and PSD) as well as synaptic glomeruli were observed in this nucleus. Among these, the PSD profiles were observed more frequently in AChE-positive portions than in AChE-negative zones. Furthermore, the number of glomeruli was significantly higher in AChE-positive than in AChE-negative zones. Following the injection of BDA into PPT, labeled terminals within LM-Sg were rather more concentrated in the AChE-positive portion. Although the majority of PPT terminals made synaptic contacts with dendrites in the neuropil, a few terminals were involved in the synaptic glomeruli. The present results show that the synaptic organization is distinctly different between the AChE-positive and AChE-negative portions of LM-Sg. These results suggest that the AChE-positive portions of LM-Sg are relatively more involved in integrating information arising from a diverse set of inputs and processing that information within glomeruli in a complex manner than occurs in the AChE-negative portion of LM-Sg.

Cellular correlates of stages of memory formation in the chick following passive avoidance training.

The process of memory formation has been investigated using the model of one-trial passive avoidance training in the one-day old domestic chick. We have unraveled a biochemically coherent cascade of processes which, beginning with transient ion and neurotransmitter flux, and by way of a sequence of interacting pre- and post-synaptic intracellular signalling steps, results in gene activation and the synthesis of cell adhesion molecules which appear to be the effective agents in the structural processes involved in remodelling of synaptic and neuronal circuits. Further, in a related series of experiments we have shown that these biochemical and morphological changes are accompanied by significant changes in the neurophysiological status of the neurons on the IMHV and LPO, in particular in terms of their engagement in bouts of high-frequency firing. However, much remains to be clarified, particularly the meaning of the time-dependent shifts in the location of the trace, and the ways in which these molecular and cellular events translate into changes in behavior in the animal.

Nigral innervation of cholinergic and glutamatergic cells in the rat mesopontine tegmentum: light and electron microscopic anterograde tracing and immunohistochemical studies.

The substantia nigra (SN) has long been known as an important source of afferents to the pedunculopontine tegmental nucleus (PPN). However, it has not been established which of the chemospecific cell populations receive this synaptic input. We sought to address this issue by a correlative light and electron microscopic approach that combines anterograde tracing of nigral efferents with pre-embedding choline acetyltransferase (ChAT) and/or glutamate (Glu) immunohistochemistry. Following large bilateral injections of Phaseolus vulgaris-leucoagglutinin (PHA-L) in the SN, the labeled nigrotegmental fibers were concentrated in a small area of the mesopontine tegmentum which contained very few ChAT-immunoreactive (ChAT-ir) cell bodies. However, strands of fine varicose fibers penetrated to adjacent regions of the PPN which harbored numerous cholinergic perikarya. The anterogradely labeled boutons were often seen in the proximity of ChAT-ir perikarya and dendrites, but the majority (82-93%) established symmetric synaptic junctions with noncholinergic profiles. In the pars dissipata of the PPN (PPNd), one-third of the labeled terminals synapsed onto noncholinergic perikarya and primary dendrites, while in the pars compacta of the PPN (PPNc) axosomatic synapses were rare. The possibility that the perikarya receiving a rich synaptic input from the SN are glutamatergic was tested in experiments combining anterograde transport of biotinylated tracers biocytin and dextran-amine (BDA) with glutamate immunohistochemistry. In double-labeled sections, Glu-ir perikarya within the terminal plexus of nigrotegmental fibers were surrounded by synaptic terminals. The PPNd also contained retrogradely BDA-labeled neurons which were contacted by anterogradely labeled terminals. These results indicate that although a small subpopulation of cholinergic neurons in the mesopontine tegmentum receive direct synaptic input from the SN, the primary target of nigrotegmental fibers are glutamatergic cells in the PPNd. Our results also provide ultrastructural evidence that some nigrotegmental fibers innervate pedunculonigral neurons.

NMDA-receptor-mediated synaptic currents in guinea pig laterodorsal tegmental neurons in vitro.

1. Whole cell voltage-clamp techniques were used to record glutamate-receptor-mediated synaptic currents from neurons of the laterodorsal tegmental nucleus (LDT). The principal cells of the LDT contain acetylcholine and nitric oxide synthase, and are believed to be involved in the control of sleep-waking behavior via widespread projections to the thalamus and brain stem. LDT cells were recorded from slices of mature guinea pig brain stem with patch pipette solutions containing cesium as the primary cation. 2. Application of N-methyl-D-aspartate (NMDA) elicited currents that were strongly voltage dependent with a mean reversal potential of +16.3 mV. Peak currents occurred near -15 mV, and a region of negative slope conductance was seen at more negative potentials. Application of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid evoked currents that exhibited a nearly linear current-voltage relation with a mean reversal potential of -3.4 mV. 3. Electrical stimulation of local afferents elicited dual-component excitatory postsynaptic currents (EPSCs) with decays that were well fitted by the sum of two exponentials. Mean decay time constants at -60 mV were 8.77 ms for the faster component and 129.4 ms for the slower component. The faster component displayed a linear current-voltage relation and was blocked by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) or 6,7-dinitroquinoxaline-2,3-dione, indicating that it was mediated by non-NMDA receptors, whereas the slower component displayed a voltage dependence similar to that for NMDA-evoked currents and was blocked by 2-amino-5-phosphonopentanoic acid (AP-5), indicating its mediation by NMDA receptors. 4. The fractional contribution of NMDA receptors to the EPSC was estimated from double-exponential curve fits to the decay phases. With this method, NMDA receptors were estimated on average to carry 10.1% of the total peak EPSC at -60 mV. Blockade of the non-NMDA-receptor-mediated component with CNQX revealed a residual EPSC whose amplitude was 14.4% of the control value, whereas AP-5 alone reduced the control EPSC peak by 16.1%, both values were comparable with those obtained from curve fit estimates. 5. Previous work has shown that the presence of 4-aminopyridine-sensitive, A-like transient current in LDT cells is correlated with the cholinergic phenotype. The majority of cells in this study exhibited A-like transient currents that were blocked by 4-amino-pyridine, suggesting that the majority of the data were obtained from the cholinergic and NOS-containing neurons of the LDT nucleus. 6. These experiments demonstrate the synaptic activation of functional NMDA and non-NMDA receptors in LDT neurons, and indicate that NMDA receptors contribute to fast excitatory transmission in these cells. The results suggest that afferents releasing excitatory amino acids may play an important role in controlling the state-dependent activity of LDT neurons.

Distinct properties of neuronal and astrocytic endopeptidase 3.4.24.16: a study on differentiation, subcellular distribution, and secretion processes.

Endopeptidase 3.4.24.16 belongs to the zinc-containing metalloprotease family and likely participates in the physiological inactivation of neurotensin. The peptidase displays distinct features in pure primary cultured neurons and astrocytes. Neuronal maturation leads to a decrease in the proportion of endopeptidase 3.4.24.16-bearing neurons and to a concomitant increase in endopeptidase 3.4.24.16 activity and mRNA content. By contrast, there is no change with time in endopeptidase 3.4.24.16 activity or content in astrocytes. Primary cultured neurons exhibit both soluble and membrane-associated endopeptidase 3.4.24.16 activity. The latter behaves as an ectopeptidase on intact plated neurons and resists treatments with 0.2% digitonin and Na2CO3. Further evidence for an association of the enzyme with plasma membranes was provided by cryoprotection experiments and electron microscopic analysis. The membrane-associated form of endopeptidase 3.4.24.16 increased during neuronal differentiation and appears to be mainly responsible for the overall augmentation of endopeptidase 3.4.24.16 activity observed during neuronal maturation. Unlike neurons, astrocytes only contain soluble endopeptidase 3.4.24.16. Astrocytes secrete the enzyme through monensin, brefeldin A, and forskolin-independent mechanisms. This indicates that endopeptidase 3.4.24.16 is not released by classical regulated or constitutive secreting processes. However, secretion is blocked at 4 degrees C and by 8 bromo cAMP and is enhanced at 42 degrees C, two properties reminiscent of that of other secreted proteins lacking a classical signal peptide. By contrast, neurons appear unable to secrete endopeptidase 3.4.24.16.

Synaptic organization of afferent projections to the supramammillary nucleus of the rat.

We studied the fine structure of afferent terminals from the preoptic area, the nucleus of the diagonal band of Broca, the infralimbic cortex and the laterodorsal tegmental nucleus within the supramammillary nucleus (SUM) using the anterograde tracing method of horseradish peroxidase conjugated with wheat germ agglutinin (WGA-HRP). Injection of WGA-HRP into the preoptic area permitted ultrastructural recognition of many anterogradely labeled terminals in the SUM. Almost all labeled terminals (99%) contained clear round synaptic vesicles and formed asymmetric synaptic contacts (Gray's type I). About 86% of labeled terminals from the nucleus of the diagonal band were asymmetric (Gray's type I), whereas 14% contained pleomorphic synaptic vesicles and formed symmetric synaptic contacts (Gray's type II). Almost all labeled terminals from the infralimbic cortex were located in the ventral part of the SUM, and 95% of labeled terminals were Gray's type I. The majority of labeled terminals (90%) from the laterodorsal tegmental nucleus were Gray's type I, and the remaining (10%) were Gray's type II. The percentage of labeled terminals with dense-cored vesicles was very high in terminals from the preoptic area (70%), and low in terminals from the infralimbic cortex (19%). Labeled terminals in all cases contacted mainly intermediate-sized dendrites (0.5-1.0 micron diameter). All cases had only a few labeled axosomatic terminals. The cases of injections into the preoptic area and the diagonal band nucleus had some reciprocal connections at the ultrastructural level.

Cholinergic, GABAergic, and glutamate-enriched inputs from the mesopontine tegmentum to the subthalamic nucleus in the rat.

In order to clarify the origin and to examine the neurochemistry and synaptology of the projection from the mesopontine tegmentum (MTg) to the subthalamic nucleus (STN), rats received discrete deposits of anterograde tracers in different regions of the MTg. Anterogradely labeled fibers were examined in the light and electron microscopes. The distribution of GABA or glutamate immunoreactivity was examined by post-embedding immunocytochemistry. The anterograde tracing demonstrated that the projection to the STN arises from at least three divisions of the MTg: the area defined by the cholinergic neurons of the pedunculopontine region (PPN-Ch 5), the more medial and largely noncholinergic midbrain extrapyramidal area (MEA) and to a lesser extent the laterodorsal tegmental nucleus (LDTg). Post-embedding immunocytochemistry revealed that there are GABA-immunopositive and immunonegative components to this projection and at least a proportion of the GABA-immunonegative component is enriched in glutamate immunoreactivity. The similarity of the morphology, trajectory and synaptology of the anterogradely labeled fibers and the choline acetyltransferase (ChAT)-immunopositive fibers supports the proposal that at least part of the projection is cholinergic. The terminals anterogradely labeled from the MTg and the ChAT-immunoreactive terminals form asymmetrical synapses with the dendrites and spines of subthalamic neurons. Both anterogradely labeled and ChAT-positive terminals make convergent synaptic contacts with GABA-immunoreactive terminals that form symmetrical synaptic contacts and are probably derived from the globus pallidus. Taken together these findings imply that the MTg sends cholinergic, GABAergic and glutamatergic projections to the STN where at least one of the functional roles is to modulate the indirect pathway of information flow through the basal ganglia that is carried via the pallidosubthalamic projection.

Midbrain paralemniscal projections to the facial nucleus: an anatomical and immunohistochemical study.

Serial 30 microns-thick sections through the midbrain tegmentum were stained with cresyl violet. The PL was found to be situated along the medial edge of the lateral lemniscus. The PL consisted of small- (10-15 microns) and medium-sized neurons (25-35 microns), and was the most prominent at the caudal level of the superior colliculus. In order to confirm the existence of the inhibitory paralemniscal-facial pathway, a combined HRP and immunohistochemical technique was use in the rat. This experiment revealed that 10.9% of the total number of GABA immunoreactive PL neurons also labeled with HRP after HRP injection was made in the medial part of the facial nucleus (FN). Electron microscopic observations were carried out on the medial part of the facial nucleus (FN) after kainic acid injection was made into the contralateral PL in the cat. The majority of degenerating PL fibers were ranged from 0.5 to 3.1 microns in diameter and made synaptic contacts with somata, proximal dendrites and dendritic profiles. These fibers, containing either round or pleomorphic vesicles, formed asymmetrical or symmetrical synapses. It was of particular interest in the present study that 40.7% of the total number of degenerating fibers make synaptic contacts with large dendrites more than 3.0 microns in diameter.

Ultrastructural relationships between terminals immunoreactive for enkephalin, GABA, or both transmitters in the rat ventral tegmental area.

The ventral tegmental area (VTA) receives extensive afferent input from neurons containing the opioid peptide enkephalin (Enk) and/or GABA. We examined the ultrastructural basis for known functional interactions between these inhibitory neuromodulators using a combined immunoperoxidase and immunogold-silver technique. As visualized with either marker in single sections, Enk-immunolabeled terminals contained numerous small clear vesicles and one or more intensely immunoreactive dense-cored vesicles. Enk-labeled terminals formed either symmetric or asymmetric synapses on small or large unlabeled dendrites. The immunoreactive dense-cored vesicles were usually detected away from these sites of synaptic contact. Terminals singly immunoreactive for GABA, or dually labeled for Enk and GABA, showed similar morphological features but formed primarily symmetric axo-dendritic synapses. In many instances, GABA- and/or Enk-immunolabeled terminals were in direct apposition to each other and formed synapses on immediately adjacent parts of a common dendrite. Close appositions were also noted between GABA- and Enk-immunoreactive axons and varicosities that did not form synapses with either common or divergent dendrites in single sections. Immunoreactive dense-cored vesicles were frequently detected at the apposed plasmalemmal surfaces between these axon terminals. The findings suggest that Enk and GABA are released from the same or convergent terminals and co-regulate the activity of common target neurons within the rat VTA. The results are also consistent with potential presynaptic interactions between these transmitters.