Comparison of vestibular and abducens internuclear projections to the medial rectus subdivision of the oculomotor nucleus in the monkey

Comparisons were made of projections from the vestibular nuclei (VN) and abducens internuclear neurons (AIN) to cell group A of the medial rectus subdivision (MRS) of the oculomotor nuclear complex. Cell group A, the major component of the MRS, receives projections only from the ipsilateral VN and the contralateral AIN. Neither ipsilateral vestibular projections to cell group A, arising from the medial vestibular nucleus, nor projections from MVN to the opposite abducens nucleus, match the massive projection of AIN to the MRS.     

Cervical afferent pathway to inferior oblique motoneuron in the cat

The neuronal pathway implicated in the vertical cervico-ocular reflex (COR) was investigated electrophysiologically in chloralose anesthetized cats. The effect of bilateral C2 dorsal root afferent stimulation on inferior oblique motoneurons (IO-MN) was investigated by intracellular recording. Control disynaptic excitatory postsynaptic potentials elicited in IO-MNs following stimulation of the contralateral anterior semicircular canal nerve (ACN) were invariably facilitated by conditioning stimulation to both ipsilateral and contralateral C2 dorsal roots (DR) in all motoneurons tested. This result indicates that inputs from the C2 DR of both sides and the contralateral ACN converge onto secondary vestibular neurons (‘common interneurons’) which project directly to the IO-MN. Common interneurons mediating vestibular and cervical excitation to the IO-MNs were studied in the vestibular nuclei on the side opposite to the motoneurons by extracellular recording. Nineteen vestibular neurons were identified as common interneurons; they were distributed in the caudal half of the lateral nucleus and the rostral half of the descending nucleus. The present experiment provides electrophysiological evidence of the projection of upper cervical afferents to the ipsilateral vestibular nuclei. The difference in neuronal organization between the horizontal and vertical COR is also briefly discussed.     

The influence of gravity on horizontal and vertical vestibulo-ocular and optokinetic reflexes in the rabbit

The influence of the linear acceleration of gravity on the vertical and horizontal vestibulo-ocular reflexes (VVOR, HVOR) as well as the vertical and horizontal optokinetic reflexes (VOKR, HOKR) has been examined in rabbits. Rabbits were mounted in a biaxial rate table in front of a rear projection tangent screen. Eye movements were measured with a light projection technique. The HVOR, VVOR, HOKR and VOKR were measured in rabbits which were maintained both prone and supine. The gain of the HVOR for the supine orientation was reduced at all frequencies tested (0.01-0.80 Hz). Similarly there was a reduction in the gain of the HOKR. By contrast, the gain of the VVOR in the supine orientation was enhanced over a lower range of frequencies (0.02-0.04 Hz) and reduced at higher frequencies (0.10-0.80 Hz). The gain of the VOKR was not reduced in the supine orientation. The range of eye positions over which compensatory eye movements occurred was restricted in the supine orientation. The altered orientation of the medio-laterally polarized hair cells of the utricular maculae with respect to gravity in the supine orientation may cause postural instability and facilitate 'righting reflexes'. A reduction in the gains of the HVOR, VVOR and HOKR caused by linear accelerations in the sagittal plane during locomotion may decrease automatic postural responses during certain movements in which these automatic postural adjustments would not necessarily be adaptive.     

The course of direct projections from the abducens nucleus to the contralateral medial rectus subdivision of the oculomotor nucleus in the cat

We have used autoradiography (tritiated leucine) to investigate the projections of a number of nuclear groups of the cat pons. Some cells of the abducens nucleus have axons that cross the midline, ascend in the opposite median longitudinal fasciculus (MLF) and synapse on the cells of the oculomotor complex which have been identified by others as those innervating the medial rectus muscle.     

Abducens internuclear and ascending tract of deiters inputs to medial rectus motoneurons in the cat oculomotor nucleus: synaptic organization

Abducens internuclear and ascending tract of Deiters (ATD) inputs to medial rectus motoneurons in the oculomotor nucleus are important for conjugate horizontal movements. In the present study, the organization of these separate populations of neurons and their synaptic connections with medial rectus motoneurons in the cat oculomotor nucleus have been examined by light and electron microscopy by using retrograde and anterograde axonal tracers. Consistent with the patterns of retrograde horseradish peroxidase labeling, the abducens internuclear projection is predominantly, if not exclusively, contralateral, whereas the ATD projection is exclusively ipsilateral, as demonstrated by anterograde autoradiographic and biocytin labeling. Both populations of synaptic endings contain spheroidal synaptic vesicles and establish synaptic contacts with modest postsynaptic densifications. In addition, ATD synaptic endings frequently are associated with subjunctional dense bodies and subsurface cisternae. The two populations of excitatory inputs differ, however, in their soma-dendritic distribution. The majority of abducens internuclear synaptic endings contact distal dendrites, whereas the majority of ATD synaptic endings contact proximal dendrites or somata. Abducens internuclear synaptic endings furthermore have a higher density of mitochondria than ATD synaptic endings. The more proximal location of ATD synaptic endings is consistent with the faster rise time and earlier reversal to polarizing currents of ATD excitatory postsynaptic potentials in comparison to those evoked by the abducens internuclear pathway as determined electrophysiologically. Given the differences in the physiologic signals conveyed by the abducens internuclear (eye velocity and eye position) and ATD (head velocity) pathways, the findings in this study suggest that the soma-dendritic stratification of the two inputs to medial rectus motoneurons may provide a means for the separate control of visuomotor and vestibular functions, respectively.     

Abducens internuclear and ascending tract of Deiters inputs to medial rectus motoneurons in the cat oculomotor nucleus: neurotransmitters.

The abducens internuclear and ascending tract of Deiters (ATD) pathways are the principal excitatory inputs to medial rectus motoneurons in the oculomotor nucleus and are related to the control of conjugate horizontal eye movements. Differences in the morphology and soma-dendritic distribution of abducens internuclear and ATD synaptic endings are correlated with known differences in the physiological properties of these independent inputs. The present study extends these observations to the ultrastructural localization of the excitatory amino acid neurotransmitters, glutamate and aspartate, using a postembedding immunogold procedure combined with the pre-embedding immunoperoxidase localization of anterogradely transported biocytin from the abducens nucleus and the ventral lateral vestibular nucleus. Consistent with their spheroidal synaptic vesicle content and the asymmetric pre/postsynaptic membrane profile, both the abducens internuclear and ATD synaptic endings are labeled with glutamate and aspartate. However, quantitative analysis of the density of colloidal gold particles associated with mitochondria versus synaptic vesicles/axoplasmic matrix reveals significant differences in the metabolic versus neurotransmitter pools of the amino acids in the two populations of synaptic endings. The findings indicate that both aspartate and glutamate, possibly co-localized, are the excitatory neurotransmitters utilized by abducens internuclear synaptic endings whose burst-tonic physiological activity conveys information related to eye position to medial rectus motoneurons. By contrast, glutamate is the excitatory neurotransmitter associated with ATD synaptic endings whose high frequency burst activity is related to head velocity.     

Internal organization of medial rectus and inferior rectus muscle neurons in the C group of the oculomotor nucleus in monkey

Mammalian extraocular muscles contain singly innervated twitch muscle fibers (SIF) and multiply innervated nontwitch muscle fibers (MIF). In monkey, MIF motoneurons lie around the periphery of oculomotor nuclei and have premotor inputs different from those of the motoneurons inside the nuclei. The most prominent MIF motoneuron group is the C group, which innervates the medial rectus (MR) and inferior rectus (IR) muscle. To explore the organization of both cell groups within the C group, we performed small injections of choleratoxin subunit B into the myotendinous junction of MR or IR in monkeys. In three animals the IR and MR myotendinous junction of one eye was injected simultaneously with different tracers (choleratoxin subunit B and wheat germ agglutinin). This revealed that both muscles were supplied by two different, nonoverlapping populations in the C group. The IR neurons lie adjacent to the dorsomedial border of the oculomotor nucleus, whereas MR neurons are located farther medially. A striking feature was the differing pattern of dendrite distribution of both cell groups. Whereas the dendrites of IR neurons spread into the supraoculomotor area bilaterally, those of the MR neurons were restricted to the ipsilateral side and sent a focused bundle dorsally to the preganglionic neurons of the Edinger‐Westphal nucleus, which are involved in the “near response.” In conclusion, MR and IR are innervated by independent neuron populations from the C group. Their dendritic branching pattern within the supraoculomotor area indicates a participation in the near response providing vergence but also reflects their differing functional roles. J. Comp. Neurol. 523:1809–1823, 2015. © 2015 Wiley Periodicals, Inc.     

GABAergic and glycinergic neurons projecting to the trigeminal motor nucleus: A double labeling study in the rat

The distribution of GABAergic and glycinergic premotor neurons projecting to the trigeminal motor nucleus (Vm) was examined in the lower brainstem of the rat by a double labeling method combining retrograde axonal tracing with immunofluorescence histochemistry. After injection of the fluorescent retrograde tracer, tetramethylrhodamine dextran amine (TRDA), into the Vm unilaterally, neurons labeled with TRDA were seen ipsilaterally in the mesencephalic trigeminal nucleus, and bilaterally in the parabrachial region, the supratrigeminal and intertrigeminal regions, the reticular formation just medial to the Vm, the principal sensory and spinal trigeminal nuclei, the pontine and medullary reticular formation, especially the parvicellular part of the medullary reticular formation, the alpha part of the gigantocellular reticular nucleus, and the medullary raphe nuclei. Some of these neurons labeled with TRDA were found to display glutamic acid decarboxylase (the enzyme involved in GABA synthesis)-like or glycine-like immunoreactivity. Such double-labeled neurons were seen mainly in the supratrigeminal region, the reticular region adjacent to the medial border of the Vm, and the dorsal part of the lateral reticular formation of the medulla oblongata; a number of them were further scattered in the intertrigeminal region, the alpha part of the gigantocellular reticular nucleus, the nucleus raphe magnus, the principal sensory trigeminal nucleus, and the interpolar subnucleus of the spinal trigeminal nucleus. These neurons were considered to be inhibitory (GABAergic or glycinergic) neurons sending their axons to motoneurons in the Vm, or to local interneurons within and around the Vm. © 1996 Wiley-Liss, Inc.     

Retractor bulbi muscle responses to oculomotor nerve and nucleus stimulation in the cat

This study demonstratesthe presence of retractor bulbi motoneurons within the oculomotor nucleus which activate muscle units within all 4 slips of the cat retractor bulbi muscle. These muscle units are mechanically different and physiologically separate from retractor bulbi muscle units innervated by the abducens nerve. The retractor bulbi muscle, then, is innervated by two separate pools of motoneurons whose axons are carried in two different cranial nerves. These observations of mechanical properties of retractor bulbi muscle suggest that the oculomotor retractor bulbi motor units may be activated during patterned eye movements.     

GABAergic and non-GABAergic projections of accessory optic nuclei, including the visual tegmental relay zone, to the nucleus of the optic tract and dorsal terminal accessory optic nucleus in rat.

This study examines the non-gamma-amino butyric acid (GABA)ergic (group I neurons) and GABAergic neurons (group II neurons) of the accessory optic system projecting to the nucleus of the optic tract (NOT)/dorsal terminal nucleus (DTN) of the accessory optic system in rat. These nuclei include the dorsal (MTNd) and ventral (MTNv) divisions of the medial terminal nucleus, the lateral terminal nucleus, the interstitial nucleus of the superior fasciculus, the posterior fibers, and the visual tegmental relay zone. GABAergic neurons of these nuclei that do not target the NOT/DTN (group III neurons) have also been observed. The fluorescent retrograde tracer fluoro-gold was injected into the pretectum, targeting the NOT/DTN and the tissue prepared immunocytochemically to reveal neurons containing the neurotransmitter GABA. Three groups of neurons (groups I, II, and III neurons) were examined in terms of their distribution, density, and percentage present. Group I neurons are single-labeled with fluoro-gold and represent non-GABAergic neurons projecting to the NOT/DTN. These neurons are of the highest density in the lateral terminal nucleus (204 neurons/mm2). Their densities are also substantial in the MTNv (120 neurons/mm2), interstitial nucleus of the superior fasciculus, posterior fibers (96 neurons/mm2), and visual tegmental relay zone (93 neurons/mm2). Group II neurons are double-labeled with fluoro-gold and GABA. They form a system of GABAergic neurons projecting to the NOT/DTN, which are exceedingly dense in the MTNd (78 neurons/mm2) but are also dense in both the visual tegmental relay zone (49 neurons/mm2) and MTNv (33 neurons/mm2). Group III neurons are GABAergic neurons that do not target the NOT/DTN but must project to other brain nuclei and/or be interneurons. These are of extremely high concentration in the visual tegmental relay zone (316 neurons/mm2) and are also of substantial densities in the MTNd (77 neurons/mm2), lateral terminal nucleus (72 neurons/mm2), and MTNv (44 neurons/mm2). The MTNd has the highest percentage of GABAergic neurons projecting to the NOT/DTN (72%). GABAergic neurons also form significant percentages of the projections to the NOT/DTN from the visual tegmental relay zone (34%) and MTNv (21%). The percentage of the total GABAergic neurons that project to the NOT/DTN is the highest in the MTNd (50%) and MTNv (42%). The described GABAergic afferents to the NOT/DTN may function to process information concerned with the compensation for retinal slip.     

Ultrastructural study of the GABAergic and cerebellar input to the nucleus reticularis tegmenti pontis

The nucleus reticularis tegmenti pontis is an intermediate of the cerebrocerebellar pathway and serves as a relay centre for sensorimotor and visual information. The central nuclei of the cerebellum provide a dense projection to the nucleus reticularis tegmenti pontis, but it is not known to what extent this projection is excitatory or inhibitory, and whether the terminals of this projection contact the neurons in the nucleus reticularis tegmenti pontis that give rise to the mossy fibre collaterals innervating the cerebellar nuclei. In the present study the nucleus reticularis tegmenti pontis of the cat was investigated at the ultrastructural level following anterograde and retrograde transport of wheat germ agglutinin coupled to horseradish peroxidase (WGA-HRP) from the cerebellar nuclei combined with postembedding GABA immunocytochemistry. The neuropil of this nucleus was found to contain many WGA-HRP labeled terminals, cell bodies and dendrites, but none of these pre- or postsynaptic structures was double labeled with GABA. The vast majority of the WGA-HRP labeled terminals contained clear spherical vesicles, showed asymmetric synapses, and contacted intermediate or distal dendrites. Many of the postsynaptic elements of the cerebellar afferents in the nucleus reticularis tegmenti pontis were retrogradely labeled with WGA-HRP, while relatively few were GABAergic. We conclude that all cerebellar terminals in the nucleus reticularis tegmenti pontis of the cat are nonGABAergic and excitatory, and that they contact predominantly neurons that project back to the cerebellum. Thus, the reciprocal circuit between the cerebellar nuclei and the nucleus reticularis tegmenti pontis appears to be well designed to function as an excitatory reverberating loop.     

The rostral dorsal cap and ventrolateral outgrowth of the rabbit inferior olive receive a GABAergic input from dorsal group Y and the ventral dentate nucleus

The dorsal cap and ventrolateral outgrowth of the inferior olive are involved in the control of eye movements. The caudal dorsal cap is predominantly involved in the horizontal optokinetic reflex; it receives most of its GABAergic input from the nucleus prepositus hypoglossi. In the present study, we determined the source of a major inhibitory input to the rostral dorsal cap and the ventrolateral outgrowth, which are the olivary subnuclei mainly involved in the “vertical” optokinetic reflexes. We studied these subnuclei in the rabbit with the use of retrograde tracing of horseradish peroxidase and anterograde tracing of wheat germ agglutinin-coupled horseradish peroxidase combined with postembedding immunocytochemistry. The ventral dentate nucleus of the cerebellum and dorsal group y project contralaterally to the rostral dorsal cap and ventrolateral outgrowth; this projection is entirely GABAergic. The terminals of this input form predominantly symmetric synapses with extraglomerular and intraglomerular dendrites; the remaining terminals are axosomatic. In addition, the dorsal cap and ventrolateral outgrowth contain significantly more crest synapses than any other olivary subnucleus. The terminals that form these crest synapses are derived from dorsal group y and/or the ventral dentate nucleus. None of the terminals in the dorsal cap or ventrolateral outgrowth was glycinergic.     

Development of the human motor-related thalamic nuclei during the first half of gestation, with special emphasis on GABAergic circuits

This study analyzed the expression of differentiation markers (Calbindin D28K: CaBP; parvalbumin: PARV; calretinin: CalR), gamma-aminobutyric acid (GABA) markers (GABA, glutamic acid decarboxylases: GAD65, GAD67; and GABA transporters: GAT1, GAT3), and other markers (neurotensin: NT, and neurofilament-specific protein: SMI32) in the human thalamus at 8-23 gestation weeks (g.w.), focusing on the motor-related nuclei. From 8-13 g.w. mainly CaBP was expressed in the cells while fiber bundles traversing the thalamus in addition to CaBP expressed all GABA markers except GAD67. CaBP and PARV expression patterns in different nuclei changed over the time course studied, whereas NT was expressed consistently along the anterior-lateral curvature of the thalamus. CalR and SMI were detectable at 23 g.w. in the ventral parts of the dorsal thalamus. Most remarkably, punctate GAD65 immunoreactivity in the neuropil was confined to the nigro- and pallidothalamic afferent receiving nuclei from 16 to about 21 g.w., overlapping with that of CaBP in some of these nuclei (subdivisions of the ventral anterior and mediodorsal nuclei) and with PARV in others (centromedian nucleus). During this period, GAD65 immunoreactivity can be considered a marker of the basal ganglia afferent receiving territory in the motor thalamus. GAD67-positive local circuit neurons were first detected at 12-13 g.w. in the thalamic nuclei outside the basal ganglia afferent receiving territory. In the ventral anterior and centromedian nuclei, GAD-containing local circuit neurons were not conspicuous even at 22-23 g.w. The cells of the reticular nucleus expressed GAD67 and PARV from 12 g.w. on starting in the lateral-posterior regions. By 23 g.w., both markers were expressed in about two-thirds of the nucleus except for its most medial-anterior part. The results imply spatially and temporally differential expression of GABA and differentiation markers in the developing human thalamus.     

A correlated light and electron microscopic immunocytochemical study of cholinergic terminals and neurons in the rat amygdaloid body with special emphasis on the basolateral amygdaloid nucleus

The cholinergic innervation of the rat basolateral amygdaloid nucleus (BL) was determined by the immunocytochemical localization of the acetylcholine biosynthetic enzyme, choline acetyltransferase (ChAT). ChAT-immunoreactive (ChAT-IR) elements were observed throughout the BL in the form of fine puncta and varicose fibers. Electron microscopy revealed that the immunoreactive puncta represented small terminals (0.3-1.2 micron), most of which formed synaptic contacts with unlabeled dendritic shafts or spines. Less frequently, ChAT-IR terminals established synaptic contacts with large neuronal cell bodies, which had all the characteristics of projection neurons as defined on the basis of axonal projections to the ventral striatum. ChAT-IR terminals were sometimes seen to form synaptic contacts with small neuronal cell bodies, including those of ChAT-IR neurons. The ChAT-IR boutons contained pleomorphic clear vesicles of varying size, and the large majority of the synapses were of the symmetric type. Small ChAT-IR neurons were observed in all parts of the BL. Although the ChAT-IR cell bodies varied widely in shape from typical fusiform to round, most had a more or less oval shape with a major diameter of 10-14 micron. Most of the ChAT-IR neurons seemed to display a radial bipolar dendritic pattern, but multipolar cells were also observed. The ChAT-IR neurons contained an indented nucleus, which was often eccentrically located and surrounded by a thin or moderately thin rim of cytoplasm. The results obtained are discussed in relation to a quasi-cortical organization of the BL.     

Parasolitary nucleus: A source of GABAergic vestibular information to the inferior olive of rat and rabbit

At least two subnuclei of the inferior olive, the β-nucleus, and the dorsomedial cell column (dmcc), contain vestibularly responsive neurons that receive a dense descending projection that uses γ-aminobutyric acid (GABA) as the transmitter. In contrast to the GABAergic innervation of other olivary subnuclei, the terminal boutons that terminate on neurons in the β-nucleus and the dorsomedial cell column remain intact after cerebellectomy, ruling out both the cerebellum and the cerebellar nuclei as afferent sources. By using both immunohistochemical as well as orthograde and retrograde tracer methods, we have identified the source of the GABAergic pathway to the β-nucleus and dmcc in both rat and rabbit. Under physiologic recording of single olivary neurons to guide electrode placement, we injected the bidirectional tracer, wheat germ agglutinin-conjugated horseradish peroxidase (WGA-HRP) into the β-nucleus and dmcc of the inferior olive. These injections retrogradely labeled neurons in the parasolitary nucleus (Psol) near the vestibular complex. Psol neurons were identified as GABAergic with an antibody to glutamic acid decarboxylase (GAD). In the rat, Psol neurons are small (5–7 μm in diameter) and number approximately 1,800. In the rabbit, they are slightly larger (6–9 μm in diameter) and number approximately 2,200. WGA-HRP injections in conjunction with GAD immunohistochemistry double labeled a high percentage of neurons in both the rat and rabbit Psol. Injection of the orthograde tracer Phaseolus vulgaris-leucoagglutinin into the area of the Psol revealed a projection from this region to both the β-nucleus and dmcc. Subtotal electrolytic lesions of this division of the Psol caused a substantial reduction in GAD-positive synaptic terminals in both the ipsilateral β-nucleus and dmcc. The location of these GABAergic neurons, bordering both the nucleus solitarius and caudal vestibular complex, emphasizes the importance of the Psol in the processing of both vestibular and autonomic information pertinent to postural control. J. Comp. Neurol. 392: 352–372, 1998. © 1998 Wiley-Liss, Inc.     

Localization of glutamic-acid-decarboxylase-immunoreactive axon terminals in the inferior olive of the rat, with special emphasis on anatomical relations between GABAergic synapses and dendrodendritic gap junctions

Immunocytochemical and electron microscopic methods were used to examine the GABAergic innervation of the inferior olivary nucleus in adult rats. This neuronal system was visualized with an antibody against glutamic acid decarboxylase (GAD, EC 4.1.1.15), the GABA-synthesizing enzyme. A GAD-positive reaction product was encountered only in short segments of preterminal axons and in axon terminals. Their relative number per unit area of neuropil was very similar in all olivary subnuclei. Despite this homogeneity in density, obvious intraregional differences existed. Some regions were strongly immunoreactive (the "c" subgroup, the beta nucleus, and the mediolateral outgrowth of the medial accessory olive), whereas others were weakly labeled (the dorsomedial cell column and the central zones of the medial accessory and principal olives). The strongly immunoreactive areas contained the largest and most intensively labeled axon terminals. Areas of weak labeling were filled with small, weakly immunoreactive nerve terminals. Thus, variations in size and in intensity of labeling create a specific pattern of GABA innervation, revealed by an almost continuous gradient between the above-mentioned extremes. The GAD-positive axon terminals established conventional synapses with dendrites (94% of the samples) or with cell bodies (6%). The vast majority of these synapses were type II (84%) and only a small proportion formed type I synaptic contacts (16%), regardless of the nature of the postsynaptic element. Immunoreactive terminals were also involved in the complex synaptic arrangements--the glomeruli, which characterize the olivary neuropil. Within these formations, olivary neurons were electrotonically coupled through dendrodendritic gap junctions. There was a constant association between GAD-positive axon terminals and small dendritic appendages linked by gap junctions. This association was revealed not only by the systematic presence of immunolabeled terminals directly apposed to the dendritic appendages but, more importantly, by the frequent presence of type II synapses straddling both elements. These synapses were in close proximity to the low-resistance pathways represented by the gap junctions. The strategic location of these GABA synapses is discussed in relation to recent findings indicating the possibility of a synaptic modulation of the electrical coupling: the release of GABA, by increasing nonjunctional membrane conductance, could shunt the coupling between olivary neurons. The functional decoupling of selected gap junctions would be responsible for the spatial organization of the olivary electrotonic coupling.     

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.     

Peptidergic and aminergic innervation of the facial nucleus of the rat with special reference to ontogenetic development

The distribution and ontogenetic development of several neuropeptides such as enkephalin, substance P, somatostatin, neuropeptide Y, and of monoamines such as serotonin and catecholamines in the facial nucleus of the rat were investigated with immunocytochemistry. The neuropeptides were concentrated in certain subnuclei. Enkephalin-immunoreactive fibers were distributed in the medial and dorsal subnuclei, substance P in the intermediate and dorsal subnuclei, somatostatin in the intermediate subnucleus, and neuropeptide Y in the dorsal subnucleus. The amines were distributed evenly throughout the nucleus. These distribution patterns suggest that peptidergic fibers are closely related to the functions of different subnuclei, while fibers containing monoamines are more basic--not specific to individual muscles. Few of these fibers were observed in the prenatal stage of the rat, but they increased markedly in number during the first postnatal week, and had established their innervation pattern by the tenth postnatal day, which coincides with the establishment of nerve-muscle innervation. The present study further showed that fibers containing serotonin are supplied mainly from the raphe nucleus, that catecholamine fibers are from neurons containing catecholamine surrounding the facial nucleus, and that fibers containing neuropeptide Y are from the lateral part of the caudal medullary reticular formation. These findings suggest that catecholamine and neuropeptide Y are not both present in the single neurons projecting to the facial nucleus.     

Parabrachial inputs to Fos-immunoreactive neurons in the lateral central nucleus of amygdala activated by hypotension: a light and electron microscopic study in the rat

Morphological features and functional implications of projections of the parabrachial nucleus to the central nucleus of the amygdala were investigated in the rat. The anatomical study was based on injections of the tracers horseradish peroxidase and biotinylated dextran amine. An extremely dense concentration of labeled fibers was found in the lateral and lateral capsular subdivisions of the central nucleus of the amygdala, originating mainly from the external lateral and ventral lateral subnuclei of the parabrachial nucleus. The parabrachial fibers exhibited the morphological characteristic of forming dense pericellular terminal arborizations. The functional implications of this pathway in cardiovascular functions were verified using Fos protein induction in response to hypotension induced by continuous intravenous administration of hydralazine-hydrochloride. In this paradigm, Fos immunoreactivity was found to be confined to the lateral and lateral capsular subdivisions of the central nucleus of the amygdala. Double immunostaining methods were used to visualize, at the electron microscopic level, terminals labeled by biotinylated dextran amine and Fos cell labeling. With this approach, we were able to confirm that Fos-immunoreactive neurons in the central nucleus of the amygdala receive axosomatic terminals from the parabrachial nucleus. The present findings point out that parabrachial inputs to the central nucleus of the amygdala play a relevant role in regulating cardiovascular function.