Cerebellar Purkinje cell projection to the peripheral vestibular organ in the frog.

Neurones located 200 to 300 microns from the surface of the auricular lobe of the frog cerebellar cortex, and identified as Purkinje cells, were activated antidromically from the eighth cranial nerve. A parallel anatomical study confirmed the existence of this projection. On the basis of these findings the existence of a cerebello-vestibular efferent system is postulated, the precise significance of which is as yet unclear. However, since Purkinje cells in other species have an inhibitory action on their target cells, the Purkinje efferent system to the vestibular organ may have an action similar to that ascribed to the olivo-cochlear bundle upon the cochlea, that is, to serve as an inhibitory control system.

Single neostriatal efferent axons in the globus pallidus: a light and electron microscopic study.

Intracellularly labeled rat neostriatal projection neurons were analyzed with both light and electron microscopy. The axons of medium spiny neurons were traced into the globus pallidus and were found to make synaptic contacts with pallidal dendrites. Despite the common somato-dendritic morphology of the neostriatal projection neurons, two different distribution patterns of efferent axons were observed, indicating the presence of functionally different medium spiny neurons in the neostriatum.

Dopaminergic regulation of striatal efferent pathways.

In the past year there has been a growing debate about the distribution of dopamine receptors in striatal efferent pathways. As is often the case, different approaches lead to different perspectives. Nevertheless, the available data can be reconciled with a model in which D1 and D2 dopamine receptors are segregated in the distal dendrites and axonal terminal fields of striatonigral and striatopallidal neurons, but intermingled in the soma and proximal dendrites.

Afferent modulation of dopamine neuron firing patterns.

In recent studies examining the modulation of dopamine (DA) cell firing patterns, particular emphasis has been placed on excitatory afferents from the prefrontal cortex and the subthalamic nucleus. A number of inconsistencies in recently published reports, however, do not support the contention that tonic activation of NMDA receptors is the sole determinate of DA neuronal firing patterns. The results of work on the basic mechanism of DA firing and the action of apamin suggest that excitatory projections to DA neurons from cholinergic and glutamatergic neurons in the tegmental pedunculopontine nucleus, and/or inhibitory GABAergic projections, are also involved in modulating DA neuron firing behavior.

Efferent projections of the subthalamic nucleus in the rat: light and electron microscopic analysis with the PHA-L method.

Efferent projections of rat subthalamic nucleus were studied by use of the axonal transport of phaseolus vulgaris-leucoagglutinin (PHA-L), and the results were analyzed with light and electron microscopes. PHA-L injections in the subthalamic nucleus (STH) resulted in heavy labeling of fiber plexus with en passant boutons and terminals in the pallidal complex, i.e., the entopeduncular nucleus (EP), the globus pallidus (GP) and the ventral pallidum (VP), and the substantia nigra pars reticulata (SNR). Labeling in GP was characterized by two distinct bands of labeled terminals oriented dorsoventrally, whereas labeling in SNR was patchy. STH efferents to the pallidum and SNR displayed a mediolateral topographic organization. With regard to dorsoventral organization, projections to GP were inverted, but those to SNR were not. There were moderate projections to the neostriatum and sparse projections to the frontal cortex, substantia innominata, substantia nigra pars compacta (SNC), pedunculopontine tegmental nucleus, ventral part of the central gray matter including the dorsal raphe nucleus, and the mesencephalic and pontine reticular formation. PHA-L injections in the zona incerta and the lateral hypothalamic area resulted in fiber and terminal labelings in many structures, including the basal forebrain, EP, SNC, and other brainstem areas that overlap with some of the terminal sites of STH projections. Ultrastructural observations of PHA-L labeled processes in GP and SNR revealed that STH terminals in both structures contained small pleomorphic vesicles and formed asymmetrical contacts. These contacts were mainly on dendritic shafts, but some were on somata. It also was observed that the myelinated axons of STH neurons lost their myelin after reaching their target areas and the synaptic boutons arose from relatively thin unmyelinated axons.

A collateral pathway to the neostriatum from corticofugal neurons of the rat sensory-motor cortex: an intracellular HRP study.

A projection from large pyramidal cells in layer V of the rat somatic sensory-motor (SSM) cortex both to the neostriatum and the brainstem was demonstrated by intracellular recording and injection of horseradish peroxidase (HRP). Layer V neurons that project to the brainstem were identified either by antidromic activation from the cerebral peduncle or by tracing the HRP-labeled axon into the internal capsule in histochemically processed sections. Intracellular responses to stimulation of the hindlimb, forelimb or mystacial pad were also examined. Five of 20 HRP-injected neurons that project to the brainstem had a fine collateral branch within the striatum. These branched corticostriatal cells respond at short latency (7--12 msec) to somatic sensory stimulation. All of the injected corticofugal neurons that had a striatal collateral were large pyramidal neurons located in layer Vb of the forelimb and head areas of SSM cortex. Branched corticofugal neurons have a rich basal dendritic field and a prominent apical dendrite that arborizes in the superficial cortical layers. Intracortical axon collaterals from the branched cells ramify in layers V and VI, and also project to the upper layers of cortex near the apical dentrite. Beyond the cortex, the main axon has no collateral branches, except for a single laterally directed branch in the neostriatum. The diameter of the striatal collateral is small (about 0.5 micrometer) compared to that of the main axon (2.0--2.5 micrometers). It is concluded that these branched cells provided a parallel input to the neostriatum and to brainstem or spinal motor centers.

Amygdaloid projections to the frontal cortex and the striatum in the rat.

Projections from the basolateral nucleus of the amygdala (BLA) to the frontal cortex and the striatum were studied by using Phaseolus vulgaris-leucoagglutinin (PHA-L) anterograde tracing technique in the rat. PHA-L injections into the rostral part of the BLA resulted in a dense labeling of fibers with boutons in the dorsal bank of the rhinal fissure and in the lateral and the medial agranular cortex. PHA-L injections into the caudal part of the BLA produced a dense labeling of fibers in the medial surface of the frontal cortex. In most of the cortical regions, labeled fibers were predominantly distributed in two bands: one in the deep part of layers I and II and the other, heavier band, in layers V and VI. PHA-L injections into the rostral BLA resulted in a dense labeling of fibers with boutons in the olfactory tubercle, the rostral and caudolateral portion of the nucleus accumbens, and a large region of the caudate-putamen. The labeled area of the caudate-putamen included the rostroventral area, the central area, and the area caudal to the anterior commissure and dorsal and lateral to the globus pallidus. PHA-L injections into the caudal BLA produced fiber labeling in the most rostromedial area of the caudate-putamen facing the lateral ventricle, the medial portion of the nucleus accumbens, and the lateral septum. In the rostroventral striatum, PHA-L-labeled fibers selectively innervated the matrix compartment that contains abundant somatostatin-immunoreactive fibers. Compartmental segregation was less clear in the caudodorsolateral caudate-putamen and in the nucleus accumbens. Electron microscopy revealed that PHA-L-labeled boutons in the striatum contained abundant, small, round vesicles. These boutons formed asymmetrical synapses with dendritic spines of striatal neurons.

The fine structure of the rat subthalamic nucleus: an electron microscopic study.

The normal ultrastructure of the rat subthalamic nucleus (STH) was studied. The STH consisted of tightly packed neurons distributed within a neuropil filled with large numbers of blood vessels and thinly myelinated fibers. The somata of STH neurons (diameters, D, between 10 and 25 micron) contained abundant organelles but had only a small amount of both smooth and rough endoplasmic reticulum. The nuclei had deeply invaginated nuclear envelopes and pale nucleoplasm with little heterochromatin. STH neurons often were tightly apposed without any intervening glial membranes. Similar appositions were also found between somata and dendrites, dendrites and dendrites, and dendrites and initial axon segments. Although puncta adhaerentia were often observed, no gap junctions were found on any of these membrane appositions. In the neuropil, the dendrites were mostly smooth and thin (D between 0.5 and 1 micron) with an occasional stubby spine or thin dendritic appendage. At least two types of axon terminals were identified. Type 1 terminals (D up to 1 micron) contained medium-sized round vesicles (D about 45 nm) and formed asymmetrical synapses. Type 2 terminals were often large (D up to 5 micron) and contained both round and slightly flattened vesicles (D up to 50 nm). The type 2 terminals frequently formed adherens junctions with their postsynaptic targets in addition to forming relatively symmetrical synaptic junctions. The remaining axon terminals included a small number of terminals with various morphological characteristics and possibly some tangentially sectioned type 1 and type 2 terminals. Therefore they have not been classified as individual types in this study. A quantitative analysis indicated that the type 1 terminals formed synapses mainly with thin dendrites whereas the type 2 terminals formed synapses mainly with somata and larger dendrites.

Morphology of the substantia nigra pars reticulata projection neurons intracellularly labeled with HRP.

The technique of intracellular recording and staining of the same neuron with horseradish peroxidase (HRP) was used to study the soma-dendritic and axonal morphology of nigrothalamic and nigrotectal cells in the rats. The nigrothalamic and nigrotectal cells were spread throughout the dorsoventral extent of the pars reticulata (SNR) and exhibited the same soma-dendritic and axonal features. Both populations consisted of medium-sized and large cells with extensive dendritic fields overlapping in all three directions. Their axons collateralized within the substantia nigra (SN) and in the mesencephalic tegmentum. The intrinsic collaterals were thin and branched partly within the dendritic field of a parent cell partly in remote regions of the SNR, and even in the pars compacta (SNC). The extrinsic branches involved thin arborizations in the rostroventral mesencephalic reticular substance and thicker descending and ascending collaterals. This material was supplemented by physiologically nonidentified HRP stained medium-sized and large neurons located in the SNR. The two kinds displayed the same extent and orientation of their dendrites but the branching patterns differed slightly. Proximal dendrites of all cells were coarse and smooth; thinner distal dendrites had varicosities and spinelike appendages. Some dendrites, specially those near the crus cerebri, terminated in dendritic thickets bearing many pleomorphic appendages. The orientation of dendritic fields varied with dorsoventral position of cells within the SNR. The most ventral region of the SNR contained neurons with dendrites oriented parallel to the crus cerebri and thus remained confined to the deepest stratum. The dendrites of cells in the central region of SNR were oriented mainly anteroposteriorly and ventrally, the ventral dendrites terminating in the ventralmost layer. Cells in the dorsolateral part of the SNR were characterized by the large dorsoventral extent of their dendrites which penetrated the entire thickness of SN. This variation in the arrangement of dendritic fields indicates that the SN is organized in three dorsoventral layers.

The morphology of intracellularly labeled rat subthalamic neurons: a light microscopic analysis.

Light microscopic analysis of rat subthalamic (STH) neurons which were intracellularly labeled with horseradish peroxidase, following the acquisition of electrophysiological data, revealed the following: (1) The somata of STH neurons were polygonal or oval with occasionally a few somatic spines. Usually three or four primary dendrites arose from the soma. Dendritic trunks tapered slightly and divided into long, thin, sparsely spined branches. Dendrites of some STH neurons extended into the cerebral peduncle. (2) Reconstruction of the dendritic field was made in three different planes. In either sagittal or frontal planes, the dendritic field was usually oval and the long axis was parallel to the main axis of STH. In the horizontal plane, the dendritic field of all neurons was polygonal. (3) The axons of all the neurons analyzed originated from the soma and were traced beyond the borders of STH, thus indicating that they were projection neurons. All the parent axons bifurcated at least once. After bifurcation, one axon branch coursed dorsolaterally within the cerebral peduncle and terminated in the globus pallidus. The other branch coursed caudally or mediocaudally and arborized in the substantia nigra. Frequently, the axon branches projecting toward the globus pallidus emitted fine axon collaterals within the entopeduncular nucleus. (4) About one-half of the analyzed STH neurons had intranuclear axon collaterals. The neurons with intranuclear collaterals had a higher dendritic tips/stems ratio than neurons without intranuclear collaterals. This observation indicated that STH neurons could be divided into two groups according to their axonal morphology. (5) The axonal terminal arborization observed in all the target sites (i.e., globus pallidus, entopeduncular nucleus, STH, and substantia nigra) were formed with varicose collateral branches which also gave rise to short filaments with beaded endings. Some of these projection neurons could therefore communicate with the target neurons in the globus pallidus, substantia nigra, entopeduncular nucleus, as well as STH through their collateral system.

Morphological organization of the globus pallidus-subthalamic nucleus system studied in organotypic cultures.

The morphological organization of the globus pallidus (GP), the subthalamic nucleus (STN), and the pallidosubthalamic projection was studied in organotypic cultures. Coronal slices from the GP, the STN, the striatum (CPu), and the cortex (Cx) were taken from the rat after postnatal days 0-2 and grown for 2 or 5-6 weeks. For analysis, immunocytochemistry against glutamate (GLU), parvalbumin (PV), and calretinin (CR) was combined with confocal microscopy. After 2 weeks in vitro, the STN showed a densely packed, homogeneous GLU-immunoreactive (ir) cell population. Pallidal GLU-ir neurons were heterogeneous, consisting of large-sized weakly GLU-ir neurons and small-sized intensively GLU-ir neurons. After 5-6 weeks in vitro, pallidal axons had radiated from numerous large-sized PV-ir cells and selectively innervated the STN, where they heavily ramified. Cultured STN neurons were not stained for PV; however, multipolar intensely PV-ir neurons were located at the border of the STN with their dendrites oriented towards the STN. Double labeling for PV and CR in both mature cultures and in the adult rat revealed that the culture CR-ir neurons from the GP, the Cpu, and from areas adjacent to the STN were different from cultured PV-ir neurons and their morphologies and distribution corresponded to that in vivo. These results demonstrate that 1) cultured CP and STN neurons display similar morphologies found in in vivo, 2) PV-ir pallidal neurons heavily and selectively innervate the STN; 3) there is a specific class of STN border neurons; and 4) in contrast to the in vivo situation, most cultured STN neurons are PV-negative.

The organization of divergent axonal projections from the midbrain raphe nuclei in the rat.

The intranuclear organization of divergently projecting neurons of the midbrain raphe in the rat was studied by using double retrograde axonal tracing. Paired injections of the tracers N-[acetyl-3H] WGA and horseradish peroxidase were made within known projection targets of the midbrain raphe (caudate-putamen, amygdala, hippocampus, substantia nigra, and locus coeruleus). After injections of either tracer in the aforementioned targets, retrograde labeled neurons were found mainly ipsilaterally and within midline portions of the dorsal raphe nucleus, its caudal B6 portion, and within the linear and superior central nuclei of the median raphe complex. There are discrete intranuclear distributions of raphe neurons that project to these forebrain and brainstem sites, and there is an overall rostrocaudal topographic order within the raphe with neurons projecting to the neostriatum, amygdala, and substantia nigra residing most rostrally and neurons projecting to the hippocampus and/or locus coeruleus occupying caudal portions of the B6 and superior central nuclei. Such distributions of projection neurons suggest the existence of an "encephalotopic" intranuclear organization within the raphe; that is, each central nervous system structure that receives midbrain raphe projections has its own unique representation within a topographically distinct portion of one or more of the raphe subgroups. These findings suggest an overall functional organization within the midbrain raphe nuclear complex whereby rostral portions are associated with the basal ganglia and related nuclei, and caudal portions relate to the limbic system. An intermediate representation of amygdala-projecting raphe neurons functionally conjoins the two. Collateralized neurons are found within complex zones of overlap in the topographically organized distributions of raphe neurons projecting to functionally related structures.

Relationship of the axonal and dendritic geometry of spiny projection neurons to the compartmental organization of the neostriatum.

Intracellular injection of HRP combined with immunocytochemistry for [Leu]enkephalin was used to demonstrate striatal spiny neuron dendritic and local axonal arborizations in the same section as enkephalin-rich patches (striosomes). Cobalt intensification of the first DAB reaction prior to the immunoperoxidase steps resulted in good contrast between the black reaction product in the intracellularly labeled cells and the brown staining for [Leu]enkephalin. Serial reconstructions of the labeled cells and nearby boundaries between the enkephalin-rich striosomes and enkephalin-poor matrix allowed the relationship between the arborizations of the labeled cells and these boundaries to be established. It was also possible to examine the relationship to compartmental boundaries of a second neuronal class consisting of large, pallidallike neurons whose somatodendritic morphology was outlined by immunoperoxidase-labeled terminals. We found that spiny projection neurons in both compartments have dendritic arbors and local axonal collaterals that are confined by compartmental boundaries. The termination or recurvature of dendrites at such boundaries suggests that the cellular basis of striatal compartmental organization is provided by this class of striatal neuron. On the other hand, large pallidumlike striatal neurons were found to have dendrites that extend across compartmental boundaries. These results support previous reports that striatal spiny projection neurons preserve the compartmental segregation of parallel striatal input-output systems, whereas other classes of striatal neurons may serve to provide limited integration between compartments.

Cholinergic and noncholinergic tegmental pedunculopontine projection neurons in rats revealed by intracellular labeling.

Morphological features of rat pedunculopontine projection neurons were investigated in in vitro preparation by using intracellular labeling with biocytin combined with choline acetyltransferase (ChAT) immunohistochemistry. These neurons were classified into two types (Type I and II), based on their electrical membrane properties: Type I had low-threshold Ca2+ spikes, and Type II had A-current. All Type I neurons (n = 17) were ChAT immunonegative (ChAT-). Type II neurons were either ChAT immunopositive (ChAT+; n = 49) or ChAT- (n = 20). In terms of topography in the tegmental pedunculopontine nucleus (PPN), Type I neurons were dispersed throughout the extent of the nucleus, whereas Type II neurons tended to be located more in the rostral and middle sections. Both Type I and II neurons consisted of small (long axis < 20 microns), medium (20-35 microns), and large (> 35 microns) cells. The small cells were round or oval; medium cells were round, triangular, or fusiform; and the large cells were primarily fusiform in shape. In terms of the soma size, there was a difference in Type I (15-38 microns) and Type II (11-50 microns) neurons, but no significant difference was found between Type II ChAT+ and ChAT- cells. Both types of neurons had three to six primary dendrites, but the dendritic field was more prominent in Type II neurons. Most of the axons originated from one of the primary dendrites, which gave off axon collaterals, some of which projected out of the nucleus. The intrinsic collaterals were thin and branched partly within the dendritic field of the parent cell. The extrinsic collaterals were thicker and could be grouped into three categories: 1) collaterals arborizing in the substantia nigra; 2) collaterals ascending mainly toward the thalamus, pretectal, and tectal area; and 3) collaterals descending toward the mesencephalic and/or pontine reticular formation. It was noted that the collaterals of both ChAT+ and ChAT-neurons were traced into the substantia nigra. There was no significant difference in antidromic latencies between Type I (m = 1.47 msec) and Type II (m = 1.36 msec) neurons following electrical stimulation of the substantia nigra.

An intracellular HRP study of the rat globus pallidus. II. Fine structural characteristics and synaptic connections of medially located large GP neurons.

In order to classify the presynaptic elements contacting the principle class of globus pallidus neurons, electron microscopic examination of serial sections made from a medially located large globus pallidus neuron, labeled with intracellular horseradish peroxidase, was undertaken. In addition, the use of labeled and light microscopically reconstructed material allowed us to quantitatively determine the distribution of each bouton type along the soma and dendrites. Six types of presynaptic terminals contacting the labeled cell have been recognized. Type 1 endings, the most numerous (84%), make symmetrical contacts on all portions of the cell, except spines, contain large pleomorphic, and a few large dense-core vesicles. Type 2 endings are filled with small spherical-to-ellipsoidal synaptic vesicles. They make asymmetrical contacts only with higher-order dendrites and account for 12% of synaptic contacts onto the labeled neuron. Type 3 endings are large, contain sparsely distributed large pleomorphic vesicles, and make two symmetrical synapses per bouton, one onto a spine head and the other onto the underlying dendritic shaft. They are infrequent (0.2%), being found only in association with dendritic spines. Type 4 endings contain large pleomorphic synaptic vesicles and no dense-core vesicles. They make symmetrical contacts with the short primary dendrites. Type 5 endings contain a mixture of small clear pleomorphic vesicles and numerous large dense-core vesicles. They contact only the cell body and the short primary dendrites, making up 20% of somatic synaptic contacts but less than 1% of contacts onto dendrites. Type 6 boutons contain oval and flattened synaptic vesicles and establish symmetrical contacts with higher-order dendritic branches and the cell body.

An intracellular HRP study of the rat globus pallidus. I. Responses and light microscopic analysis.

A study of the intracellularly recorded responses of rat globus pallidus neurons to activation of striopallidal fibers was combined with light microscopic examination of the morphology of these same neurons using intracellular horseradish peroxidase. The response to stimulation of caudate-putamen is an inhibitory postsynaptic potential with observed latencies ranging from 5.1 to 9.8 msec. These values correspond to conduction velocities of 0.4 to 0.8 m/second for striopallidal fibers. Comparison with extracellular controls shows no excitatory component to the response. All recovered and analyzed neurons (n = 11) were of the large type of pallidal neuron known from Golgi studies but the addition that two subtypes could be recognized. Large neurons located medially in the nucleus had dendritic fields with large dorsoventral extent (ca. 1 mm) when compared to their mediolateral and rostrocaudal dimensions (ca. 0.4 mm) and these neurons emitted no axon collaterals. Large neurons located laterally in the nucleus had disklike dendritic fields with both dorsoventral and rostrocaudal dimensions being on the order of 1 mm but with a minor axis of approximately 100 micrometers. The axons of these neurons possessed collaterals. As a consequence of their disk-shaped dendritic field, neurons belonging to the laterally placed subgroup and occupying the narrow (ca. 100 micrometers thick) striopallidal border zone known to receive a distinct input from neostriatum have dendrites restricted to that zone.

A Golgi study of rat neostriatal neurons: light microscopic analysis.

At least two types of large neurons (somatic cross-sectional areas, SA greater than 300 microns2) and five-types of medium neurons (SA between 100 and 300 microns2) were distinguished in Golgi preparations of the adult rat neostriatum. Type I large cells had aspinous somata with long, radiating, sparsely spined dendrites which were sometimes varicose distally, whereas type II large cells had spines on both somatic and dendritic surfaces. Type I medium cells had aspinous somata and proximal dendrites, but their distal dendrites were densely covered with spines. Type II medium cells had somatic spines, and their radiating dendrites were sparsely spined. Other medium cells had no somatic spines: Type III cells had poorly branched and sparsely spined dendrites. Type IV cells had profusely branched, sparsely spined dendrites. Type V cells had radiating and varicose dendrites which could also be sparsely spined. Several small neurons (SA mostly less than 100 microns2) were also found in the rat neostriatum: Some had aspinous soma with sparsely spined dendrites; others had somatic spines. Except for the type II large cells, intrinsic axon collaterals were observed for every type of neuron, indicating that they all had local integrating functions.

Morphological and electrophysiological characteristics of projection neurons in the nucleus interpositus of the cat cerebellum.

The populations of neurons in the nucleus interpositus (IP) of the cat cerebellum which project to the ventral lateral nucleus of the thalamus (VL), the red nucleus (RN), the nucleus reticularis tegmenti pontis (NRTP), the pontine nuclei (PN), the inferior olive (IO), and the cerebellar cortex were identified by intracellular and extracellular injections of HRP and studied electrophysiologically. When HRP was simultaneously injected into the VL, RN, and IO, over 95% of the neurons in the IP nuclei were labeled; indicating that there are few, if any, local circuit neurons. The vast majority (86%) of the larger IP neurons (soma length greater than or equal to 20 micrometer) project rostrally to the RN and thalamus. These neurons typically have long, relatively spine free dendrites and axons which in a few cases gave rise to recurrent collaterals. Two intracellularly stained projection neurons which had exceptionally long spiny dendrites had axons which gave rise to nucleocortical collaterals in addition to several local collaterals. IP neurons projecting to the NRTP and PN were located primarily in the lateral aspect of the nucleus interpositus anterior. Electrophysiological experiments established that neurons projecting to the NRTP also project to the VL. The IP neurons projecting to the IO have small fusiform or multipolar somata, long thin dendrites, and receive excitatory inputs from the IO. At least 73% of the small neurons in the IP project to the IO, and some of these, in addition, project to the VL. There are at least three morphologically distinguishable populations of projection neurons, small IO projections neurons, and neurons with nucleocortical collaterals. The projection of the IP to diverse regions of the brain is accomplished mainly by axon collateralization, but regional and morphological specialization also play a role in the organization of the output of the IP.

An HRP and autoradiographic study of the projection from the cerebellar cortex to the nucleus interpositus anterior and nucleus interpositus posterior of the cat.

The recently developed anatomical techniques of retrograde transport of the enzyme horseradish peroxidase (HRP), anterograde transport of tritiated amino acid, and intracellular injections of HRP were used to study the organization of the corticonuclear projection to the nucleus interpositus anterior (NIA) and the nucleus interpositus posterior (NIP) of the cat. Injections of HRP into the NIA and the NIP revealed that the major areas of the cortex which provided afferents to these two nuclei were the intermediate cortex of the anterior lobe (IAL) and the paramedian lobule (PML). There were, however, significant differences in the distribution of Purkinje (Pk) cells which projected to each nucleus. The NIA received afferents from all areas of the IAL while the NIP projection area was restricted to a band located at the medi-almost aspect of the lobe. All areas of the PML, in particular the intermediate folia, projected to the NIP, while the Pk cells which sent axons to the NIA were restricted to the rostral and caudal folia of this lobule. The projection from each area was somatotopically organized. The axons of intracellularly stained Pk cells were followed to their termination in the NIA and NIP confirming the results obtained with the two extracellular techniques. An attempt was made to examine the organization of the corticonuclear projection at the single cell level in the PML. Pk cells located in the same sagittal plane appeared to terminate in the same area of the same nucleus while Pk cells located not more than 500 micrometers medial or lateral to each other terminated in different nuclei. Basically, the organization of the corticonuclear projection from the IAL is longitudinally organized while the PML has a much more complex arrangement in which the Pk cells projecting to the NIA and NIP are interspersed.

Ionic mechanisms involved in the spontaneous firing of tegmental pedunculopontine nucleus neurons of the rat.

We have previously defined three types of tegmental pedunculopontine nuclei neurons based on their electrophysiological characteristics: Type I neurons characterized by low-threshold Ca2+ spikes, Type II neurons which displayed a transient outward current (A-current), and Type III neurons having neither low-threshold spikes nor A-current [Kang Y. and Kitai S. T. (1990) Brain Res. 535, 79-95]. In this report, ionic mechanisms underlying repetitive firing of Type I (n=15) and Type II (n=69) neurons were studied in in vitro slice preparations. Type I neurons did not fire rhythmically but their spontaneous firing frequency ranged from 0 to 19.5 spikes/s (mean 9.7 spikes/s). The spontaneous firing of Type II neurons was rhythmic, with a mean frequency of 9.6 spikes/s (range 3.5-16.0 spikes/s). Choline acetyltransferase immunohistochemistry combined with biocytin labeling indicated that none of the Type I neurons were immunopositive to choline acetyltransferase, while 60% (42 of 69) of Type II neurons were immunopositive. There was no apparent difference in the electrophysiological membrane properties of immunopositive and immunonegative Type II neurons. At membrane potentials subthreshold for Na+ spikes (-50 mV), spontaneous membrane oscillations (11.6 Hz) were observed: these underlie the spontaneous repetitive firing of Type I neurons. The subthreshold membrane oscillation was tetrodotoxin sensitive but was not affected by Ca2+-free medium. A similar tetrodotoxin-sensitive subthreshold membrane oscillation (10.5 Hz) was also observed in Type II neurons. However, in Type II neurons a membrane oscillation was also observed at higher membrane potentials (-50 mV). This high-threshold oscillation was insensitive to tetrodotoxin and Na+-free medium, but was eliminated in Ca2+-free conditions. The amplitude and frequency of the high-threshold oscillation was increased upon membrane depolarization. At the most prominent oscillatory level (around -40 mV), the high-threshold oscillation had a mean frequency of 8.8 Hz. The high-threshold Ca2+ spike was triggered from the peak potential (-35 to -30mV) of the high-threshold oscillation. Application of tetraethylammonium chloride (< 5 mM) increased the amplitude of the high-threshold oscillation, while nifedipine greatly attenuated the high-threshold oscillation without changing the shape of the high-threshold Ca2+ spike. Application of Cd2+ eliminated both the high-threshold oscillation and the high-threshold Ca2+ spike, and omega-conotoxin reduced the size of the high-threshold Ca2+ spike without affecting the frequency of the high-threshold oscillation. Nickel did not have any effect on either the high-threshold oscillation or the high-threshold Ca2+ spike. These data suggest an involvement of N- and L-type Ca2+ channels in the generation of the high-threshold oscillation and the high-threshold Ca2+ spike, respectively. The results indicate that a persistent Na+ conductance plays a crucial role in the subthreshold membrane oscillation, which underlies spontaneous repetitive firing in Type I neurons. On the other hand, in addition to a persistent Na+ conductance for subthreshold membrane oscillation, a voltage-dependent Ca2+ conductance with Ca2+-dependent K+ conductance (for the high-threshold oscillation) may be responsible for rhythmic firing of Type II neurons.