Injections Of Wheat Germ Agglutinin Research Articles

The Location of Spinothalamic Axons within Spinal Cord White Matter in Cat and Squirrel Monkey

The locations of spinothalamic (STT) fibers in the spinal cord white matter have been identified in cat and squirrel monkey by light-microscopic visualization of labeled fibers following multiple thalamic injections of wheatgerm agglutinin conjugated to horseradish peroxidase. Thalamic injections were combined with either a constricting dural tie or an intraspinal injection of colchicine to facilitate axonal labeling at more rostral spinal levels. In the cat, the ventral-to-dorsal distribution of labeled STT fibers was bimodal. In the ventrolateral white matter, labeled axons were coarse in nature and were primarily concentrated peripherally. In the dorsolateral white matter, labeled STT axons consisted of fine-caliber fibers concentrated in the ventral portion of the dorsolateral funiculus and were equally distributed throughout the medial and lateral white matter. In the squirrel monkey, the distribution of STT fibers was unimodal, extending from the ventral surface of the spinal white matter to the ventralmost portion of the dorsolateral funiculus. As in the cat, however, the ventrally located axons were large and coarse and were primarily located in the peripheral white matter, whereas the dorsalmost STT fibers were of fine caliber and were distributed equally in the medial and lateral white matter.

Cytoplasmic transport of ribosomal subunits microinjected into the Xenopus laevis oocyte nucleus: a generalized, facilitated process.

To study the biochemistry of ribonucleoprotein export from the nucleus, we characterized an in vivo assay in which the cytoplasmic appearance of radiolabeled ribosomal subunits was monitored after their microinjection into Xenopus oocyte nuclei. Denaturing gel electrophoresis and sucrose density gradient sedimentation demonstrated that injected subunits were transported intact. Consistent with the usual subcellular distribution of ribosomes, transport was unidirectional, as subunits injected into the cytoplasm did not enter the nucleus. Transport displayed properties characteristic of a facilitated, energy-dependent process; the rate of export was saturable and transport was completely inhibited either by lowering the temperature or by depleting nuclei of ATP; the effect of lowered temperature was completely reversible. Transport of injected subunits was likely a process associated with the nuclear pore complex, since export was also inhibited by prior or simultaneous injection of wheat germ agglutinin, a lectin known to inhibit active nuclear transport by binding to N-acetyl glucosamine-containing glycoproteins present in the NPC (Hart, G. W., R. S. Haltiwanger, G. D. Holt, and W. G. Kelly. 1989. Annu. Rev. Biochem. 58:841-874). Although GlcNAc modified proteins exist on both the nuclear and cytoplasmic sides of the nuclear pore complex, ribosomal subunit export was inhibited only when wheat germ agglutinin was injected into the nucleus. Finally, we found that ribosomal subunits from yeast and Escherichia coli were efficiently exported from Xenopus oocyte nuclei, suggesting that export of some RNP complexes may be directed by a collective biochemical property rather than by specific macromolecular primary sequences or structures.

The distribution and morphology of corticogeniculate axons in ferrets

This study examined the distribution of axons in the lateral geniculate (LGN) and adjacent nuclei following injections of wheat germ agglutinin bound to horseradish peroxidase (WGA-HRP), [ 3H]proline and Biocytin into area 17 in ferrets. Terminal label was found in the LGN, perigeniculate (PGN), medial interlaminar (MIN) and lateral posterior (LP) nuclei. LGN label was more concentrated in the interlaminar zones. Analysis of axonal morphology showed that different axon types project to LGN, MIN and LP. These findings suggest that feedback pathways from area 17 are complex and composed of several classes of axons.

Lumbar dorsal root projections to spinocerebellar cell groups in the rat spinal cord: a double labeling study

The aim of this study has been to investigate projections to spinocerebellar cell groups from lumbar dorsal root ganglia (DRGs) in the rat. The binding subunit of cholera toxin conjugated to horseradish peroxidase (B-HRP) was used to label primary afferent fibers. Spinocerebellar neurons were labeled retrogradely by Fluoro-Gold (FG). To determine the orientation of dendrites, retrogradely labeled spinocerebellar neurons were studied, following injections of wheat germ agglutinin conjugated horseradish peroxidase (WGA-HRP) into cerebellum. FG or WGA-HRP labeled neurons were found mainly in laminae V and VII, in the lateral group of lamina IX, in Clarke's column (CC) and in the dorsal funiculus. B-HRP labeled primary afferent fibers overlapping with FG labeled cells were observed at all these locations after injections of B-HRP into different DRGs. The overlap in lamina V was found mainly medially and dorsolaterally. CC was found to receive dense projections from DRGs L1-6. In the lumbar part of CC, labeling from DRGs L4-5 overlapped and was distributed over the entire mediolateral extent of the CC, whereas labeling from DRGs L1-3 was somatotopically organized and projected to successively more dorsomedial areas. The central area of lamina VII showed moderate labeling from DRGs L3-5. The lateral group of lamina IX received only smaller amounts of labeled fibers from DRGs L3-5.

Anterograde transsynaptic transport of WGA-HRP from spinal afferents to postganglionic sympathetic cells of the stellate ganglion of the guinea pig

After injection of wheat germ agglutinin conjugated horseradish peroxidase (WGA-HRP) or choleragenoid conjugated HRP (B-HRP) into lower cervical and upper thoracic dorsal root ganglia (DRG), HRP reaction product was observed in peripheral fibers of spinal afferents and in postganglionic cell bodies of the stellate ganglion (SG) in the guinea pig. After WGA-HRP injection into C8-T3 or T5DRG, HRP-labelled cells were observed to cluster at the rami within the SG, with peak labelling observed 36 h after injection. SG cell labelling occurred with B-HRP as well, but not with native HRP after injection into thoracic DRG. Injection of this tracer in C8DRG gave rise to a small number of labelled cells. In contrast to the labelling pattern following thoracic or C8DRG injections, injection of WGA-HRP or native HRP into C6DRG, led to random SG cell labelling. We conclude that the anterograde transsynaptic transport, following injection of WGA-HRP into thoracic DRG, provides a method to selectively label a population of postganglionic sympathetic neurons within the SG. A combination of transsynaptic and retrograde transport appears to be responsible for labelling after injection into C8DRG, whereas labelling after C6DRG injections seems to be due primarily to retrograde transport.

Relationships between patterns of acetylcholinesterase activity and geniculocortical terminal fields in developing and mature rat visual cortex

Intraocular injections of wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP) result in anterograde transneuronal labeling of geniculocortical axon terminals in cortical area 17. In area 17 of developing rat pups, transported WGA-HRP occurs primarily in layer I and in a band that includes layer IV and deep layer III; this pattern is virtually identical to the laminar pattern of endogenous acetylcholinesterase (AChE) activity. In adult animals, transported WGA-HRP again is localized in layer I and in deep layer III and layer IV, but the endogenous AChE activity is found most prominently in deep layer IV and layer V. These results indicate that geniculocortical terminal fields are co-extensive with transient patterns of AChE activity in the developing rat, but not with the mature pattern of AChE in the adult.

Combined Retrograde Tracing and Immunocytochemical Identification of Luteinizing Hormone-Releasing Hormone- and Somatostatin-Containing Neurons Projecting to the Median Eminence of the Rat*

LHRH and somatostatin or somatotropin-release inhibiting factor (SRIF) are produced by neurons whose cell bodies are located in telencephalic and diencephalic regions in the rat. Many, but not all, of these neurons project to the external zone of the median eminence (ME), where the peptides are released from the nerve terminals into hypophysial portal vessels. In the present study, we identified these neurons by in vivo injection of a retrograde tracer, the lectin wheat germ agglutinin (WGA), into the external zone of the ME. Subsequently, colchicine was given into the lateral ventricle 10-24 h after the WGA injection. The animals were killed 24-48 h after the WGA injection. Vibratome sections of the brains were stained for both WGA and LHRH or SRIF with a dual immunocytochemical technique. Approximately 70% of the LHRH neurons in the septum and the anterior hypothalamus and about 70% of the SRIF neurons in the medial preoptic area, the anterior periventricular area, and the paraventricular nucleus were double labeled, indicating that they projected to the ME. None of the SRIF neurons in the ventromedial and arcuate nuclei were labeled with WGA. Double labeled LHRH cells were either smooth and fusiform or spiny. WGA-accumulating LHRH or SRIF perikarya were intermixed with single labeled LHRH or SRIF cells, which apparently did not project to the ME. The results indicate that there are at least two populations of LHRH neurons in the preoptic-septal region and two populations of SRIF neurons in the medial preoptic and anterior periventricular areas and the paraventricular nucleus of the rat brain: one with access to the portal capillaries of the ME and, therefore, functionally related to the regulation of the pituitary, and another without access to portal capillaries, perhaps functionally related to intracerebral neurotransmission or modulation. Moreover, some hypophysiotropic LHRH and SRIF neurons may have axon collaterals reaching multiple targets within the central nervous system.

Cerebellar connections with the motor cortex and the arcuate premotor area: An analysis employing retrograde transneuronal transport of WGA‐HRP

We have employed transneuronal transport to examine the anatomical relationships between the deep cerebellar nuclei and 2 cortical motor areas: the primary motor cortex and the arcuate premotor area (APA). In the same animals, we have also examined the patterns of labeling in the thalamus and the red nucleus to provide evidence for the potential routes of transneuronal transport to the cerebellum. When the appropriate technical procedures were employed, cortical injections of wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP) resulted in transneuronal labeling within portions of the contralateral deep cerebellar nuclei. Injections into the primary motor cortex labeled neurons in the dentate and in the 2 subdivisions of the interpositus. Injections into the APA labeled neurons in the dentate and in only the posterior subdivision of the interpositus. In most instances, dentate neurons were more intensely labeled following the cortical injections than interpositus neurons. The transneuronal labeling observed in the dentate nucleus was topographically organized. The dentate region that was labeled following injections into the "arm area" of the APA was caudal and ventral to the dentate region that was labeled following injections into the "arm area" of the primary motor cortex. This observation provides evidence for two "arm areas" in the dentate: one anatomically related to the APA, and the other related to the primary motor cortex. More than one route of transport may be responsible for the labeling of cerebellar neurons. We propose that the labeling observed in the dentate nucleus reflects the pattern of connections in the cerebellothalamocortical pathways that link the dentate with the cerebral cortex. Thus, our observations support the concept proposed by Schell and Strick (J. Neurosci. 4:539-560, '84)--that the cortical targets of the dentate nucleus include both the primary motor cortex and the APA.

Cholinergic innervation of the superior colliculus in the cat

The superficial and intermediate gray layers of the superior colliculus are heavily innervated by fibers that utilize the neurotransmitter acetylcholine. The distribution, ultrastructure, and sources of the cholinergic innervation of these layers have been examined in the cat by using a combination of immuno-cytochemical and axonal transport methods. Putative cholinergic fibers and cells were localized by means of a monoclonal antibody to choline acetyltransferase (ChAT). ChAT immunoreactive fibers are distributed throughout the depth of the superior colliculus, with particularly dense zones of innervation in the upper part of the superficial grey layer and in the intermediate grey layer. Within the superficial grey layer, the fibers form a continuous, dense band, whereas within the intermediate grey layer the fibers are arranged in clusters or patches. Although the patches are present throughout the rostrocaudal extent of the superior colliculus, they are most prominent in middle to caudal sections. The structure of the ChAT immunoreactive terminals was examined electron microscopically. The appearance of the terminals is similar in the superficial and intermediate grey layers. They contain closely packed, mostly round vesicles, and form contacts with medium-sized dendrites that exhibit small, but prominent postsynaptic densities; a few of the terminals contact vesicle-containing profiles. To identify the sources of the cholinergic input to the superior colliculus, injections of wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP) were made in the superior colliculus and the sections were processed to demonstrate both the retrograde transport of WGA-HRP and ChAT immunoreactivity. Neurons containing both labels were found in the parabigeminal nucleus, and in the lateral dorsal and pedunculopontine tegmental nuclei of the pontomesencephalic reticular formation. Almost every cell in these nuclei that contained retrograde label was also immunoreactive for ChAT. The similarities between the laminar distributions of the ChAT terminals and the terminations of the pathway from the parabigeminal nucleus (Graybiel: Brain Res. 145:365-374, '78) support the view that the latter nucleus is a source of the cholinergic fibers in the superficial grey layer. The possibility that the pedunculopontine tegmental nucleus is a source of cholinergic fibers in the deep layers was tested by examining the distribution of labeled fibers following injections of WGA-HRP into this region of the tegmentum. Patches of labeled terminals were found in the intermediate grey layer that resemble in distribution the patches of ChAT immunoreactive fibers in this layer.(ABSTRACT TRUNCATED AT 400 WORDS)

Retinogeniculate projection fibers in the monkey optic nerve: A demonstration of the fiber pathways by retrograde axonal transport of WGA‐HRP

Seven Japanese monkeys (Macaca fuscata) were used to investigate the fiber pathways of the optic nerve. Optic nerve fibers and retinal ganglion cells were retrogradely labeled by iontophoretic injections of wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP) into electrophysiologically defined positions of the lateral geniculate nucleus (LGN). By gross anatomical observation, the optic nerve usually had one distinct bend, which flexed dorsally 3-4 mm from the eyeball, and occasionally another ventrally directed bend was found just behind the eyeball. In the optic nerve head, fibers from the various retinal areas were arranged in a wedge according to the fiber trajectory on the retinal surface. For about a 3 mm distance from the disc, fibers rapidly spread out radially. Subsequently, rather than scattering dorsoventrally, they progressed to the chiasm with a gradual increase in the degree of mediolateral (nasotemporal) scatter. The degree of the scatter was different depending on the retinal site from which the axons originated. Fibers from the peripheral retina spread out widely for a few millimeters behind the eyeball. Thereafter the scatter was rather limited until the chiasm. On the other hand, the scatter of fibers from the foveal and parafoveal areas progressed gradually through the nerve. The present study also suggests that the difference in scatter depends on the types of cells of origin. Fibers from large ganglion cells displayed more extensive scatter than fibers from medium-sized cells. In spite of the extensive scatter of fibers, two clear segregations were found; one was a dorsoventral segregation, which was displayed by both central and peripheral retinal fibers, and the other was a center-peripheral segregation in which the fibers from the nasal central (papillomacular) retina were located almost exclusively in the central part of the optic nerve surrounded by peripheral retinal fibers. However, the temporal central retinal fibers were located in the lateral periphery of the nerve, and they overlapped significantly with fibers from the temporal peripheral retina. Furthermore, a broad intermingling was found between nasal and temporal peripheral retinal fibers owing to their mediolateral scatter. Thus, the present findings based on more precise anatomical techniques indicate that the classical notion of the retinal quadrant topography in the monkey optic nerve probably is suspect. In addition, the "rotation" of the fiber arrangement was not demonstrated.

Amygdaloid projections to commissural interneurons for masticatory motoneurons

Injections of wheat germ agglutinin conjugated horse-radish peroxidase (WGA-HRP) into the trigeminal motor nucleus resulted in retrograde labeling of neurons, called commissural interneurons for masticatory motoneurons, in the contralateral supratrigeminal region. Further HRP studies, in which WGA-HRP injections were made into the amygdala and supratrigeminal region, indicated that the supratrigeminal region receives fibers from the central nucleus of the amygdala ipsilaterally. These findings raised the possibility of direct connections between the amygdala and commissural interneurons. In order to confirm the connections, electrolytic lesions in the central nucleus of the amygdala and WGA-HRP injections into the contralateral trigeminal motor nucleus were made on the same animal and electron microscopic observation was carried out on the supratrigeminal region. Of particular interest was that degenerating amygdalotegmental fibers synapsed upon HRP-labeled neurons.

Origin of sympathetic and sensory innervation of the elbow joint in the rat: A retrograde axonal tracing study with wheat germ agglutinin conjugated horseradish peroxidase

After injection of wheat germ agglutinin conjugated horseradish peroxidase (WGA-HRP) into the elbow joint of adult rats, labeled neurons were found in the stellate and the T2-T4 ganglia of the ipsilateral sympathetic trunk, and also in dorsal root ganglia at the C4-T4 levels. Most labeled sympathetic cells, 90% or more, were located in the stellate ganglion. The sensory innervation to the joint originated mainly from the dorsal root ganglia at the levels of C7-T1.

Visual pretectal neurons projecting to the dorsal lateral geniculate nucleus and pulvinar nucleus in the cat

We observed morphological subtypes of visual pretectal neurons ascending to the dorsal thalamus, following injections of wheat germ agglutinin conjugated to horseradish peroxidase into the dorsal lateral geniculate nucleus (LGNd) or the pulvinar nucleus. These neurons are composed of fusiform cells and small-sized multipolar cells in the olivary pretectal nucleus, superficial horizontal cells, fusiform cells, small-, medium- and large-sized multipolar cells in the optic tract nucleus and small- and medium-sized multipolar cells in the posterior pretectal nucleus. When somal size of the neurons projecting to the LGNd was compared to the size of neurons projecting to the pulvinar, the neuronal groups were not identical.

Connections of the primary auditory cortex in the common marmoset, Callithrix jacchus jacchus.

The afferent and efferent connections of the primary auditory cortex (AI) of common marmosets were traced following small injections of wheat germ agglutinin conjugated with horseradish peroxidase (WGA-HRP) made at best frequency (BF)-defined sites in the AI. After the injections the animals remained anesthetized for 15-23 hours; they were then perfused transcardially with fixative and the brains were processed for WGA-HRP reaction product. Examination of the disposition of labelled material revealed the following results. First, patches of terminal labelling, and to a lesser extent retrograde labelling, were found outside the injection site in the ipsilateral cortex rostral and caudal to the AI. Second, the region of the contralateral cortex corresponding to the injection site contained labelled terminals throughout the depth of the cortex; labelled neurons were found in the middle layers. Third, in each experiment a discrete region of the medial geniculate body (MG) contained retrogradely labelled neurons interspersed with anterogradely labelled terminals. These regions had a banded appearance, were found in the dorsal and rostral half of the MG, and shifted in location progressively dorsalward as the injection site BF increased. The presence of projection zones rostral and caudal to the AI of marmosets, and the disposition of the MG sources of projection in relation to BF, are similar to observations made on other New World monkeys. The ipsilateral corticocortical projections confirm electrophysiological evidence suggesting the existence of auditory fields rostral and caudal to the AI. The thalamocortical auditory system of the marmoset appears relatively simple, with a comparatively undifferentiated MG projecting to a cortical auditory system dominated by a large AI.

Intravitreal kainic acid severely reduces the size of the developing optic tectum in newly hatched chickens

Following a single intravitreal injection of 200 nmol of kainic acid (KA) to newly hatched chickens, there are acute and long-term effects on retinal ganglion cells in the chicken retina. Thirty min after injection, most ganglion cells showed cytoplasmic vacuolization. However, 14 days later, most ganglion cell soma appeared normal. Almost 60% of the cells in the ganglion cell layer (GCL) were lost, suggesting that displaced amacrine cells and not more than 40% of the ganglion cells had been eliminated. Following intravitreal injection of wheat germ agglutinin conjugated to horseradish peroxidase 14 days after the KA lesion, the amount of HRP reaction product was reduced to all retinorecipient layers, especially layers IIc and IId, of the tectum contralateral to the KA-treated eye. Fourteen days after the injection of kainic acid, during which the control tecta grow appreciably, all the superficial layers of the tectum contralateral to the kainic acid-lesioned eye, especially layers IIc and IId, were smaller than in controls, and did not differ in size from those seen ? contralateral to cut optic nerves. It is not clear whether this is a result of a developmental failure, or a shrinkage, or a combination of these factors. These results suggest that subtypes of ganglion cells may have a disproportionate influence in the maintenance of the cytoarchitectural integrity in the optic tectum. Alternatively the removal of the OFF-bipolar cells and amacrine cells presynaptic to ganglion cells may decrease their metabolism, and restrict the supply of trophic influences to the developing tectal cells.

Reversible inhibition of protein import into the nucleus by wheat germ agglutinin injected into cultured cells

The importance of glycoproteins located in the nuclear envelope in nuclear transport was tested by microinjection of karyophilic proteins into the cytoplasm of cultured human cells together with various lectins. Wheat germ agglutinin (WGA) blocked the nuclear transport of nucleoplasmin, a nuclear protein of Xenopus laevis oocytes, and of nonnuclear proteins conjugated with a synthetic peptide containing the nuclear localization signal sequence for simian virus 40 (SV40) large T antigen. Its inhibitory activity persisted for about 1 h after its injection into the cells and then gradually decreased. Export of at least some kinds of RNA from the nucleus seemed not to be affected by WGA even when import of the proteins into the nucleus was completely blocked (within 1 h after WGA injection). Moreover, WGA did not inhibit the passive diffusion of fluorescein isothiocyanate (FITC)-dextran (average M r, 17,900) into the nucleus. Wistaria floribunda agglutinin (WFA), concanavalin A (Con A), and lentil lectin did not block nuclear transport. These results indicate that WGA specifically blocks active protein import, but not passive diffusion of materials into the nucleus.

An extensive intrinsic association fiber system within cat area 7 revealed by anterograde and retrograde axon tracing methods.

The associational connections within area 7 on the crown of the middle suprasylvian gyrus in adult cats were investigated with extracellular axon tracing techniques. Injections of wheat germ agglutinin conjugated to horseradish peroxidase or of 3H-proline into area 7 revealed the presence of widespread intrinsic connections that extend over the rostral to caudal extent of the middle suprasylvian gyrus (up to 10 mm in one direction from injection sites). The connections have a complex organization with cell bodies and particulate label concentrated in patches. The patches are irregular in shape and do not form any obvious pattern. Although the transported label is concentrated in patches, large numbers of retrogradely labeled cells and significant quantities of anterogradely transported label are found in the spaces between patches. The long associational connections originate from cells in layers 2-6 and project to all cortical layers. Pyramidal cells are the primary source of the projections.

Accuracy of reinnervation of rat internal intercostal muscles by their own segmental nerves

The positions of internal intercostal motoneurons within their motor pool were studied, following reinnervation of the intercostal muscles by their original nerves. Six to 9 weeks after proximal nerve section in 10-d-old and adult rats, 0.1 microliter injections of wheat germ agglutinin (WGA)-HRP were made in the distal part of the reinnervated internal intercostal muscle. The corresponding region of the contralateral control muscle was also injected. The positions of the retrogradely labeled motoneurons were mapped in 100 microns transverse sections of thoracic spinal cord that had been stained for HRP according to the method of Mesulam (1982). In normal rats, motoneurons innervating distal muscle fibers are found largely in the more dorsal part of the internal intercostal motoneuron pool (Hardman and Brown, 1985). In adult rats, regenerated motor axons did not show any selectivity; distal muscle fibers were innervated by motoneurons whose cell bodies were distributed throughout the internal intercostal pool. However, in rats operated on at 10 d of age, distal intercostal muscle fibers were reinnervated by motoneurons that were distributed mainly in the dorsal part of the motor pool. These results support the view that positional signals may be of importance in organizing the distribution of axon terminals within muscles during development.

Location and connections of visual cortical areas in the cat's suprasylvian sulcus

The initial aim of the experiments described here was to identify and quantify the cortical and thalamic connections of visual cortical areas located in the vicinity of the suprasylvian sulcus. Inputs to various sites in this region were studied by making small injections of wheat germ agglutinin (conjugated to horseradish peroxidase) at physiologically identified locations. Retrogradely labeled cells were counted in each identifiable area of cortex and in thalamic nuclei. Some injections yielded quantitatively similar distributions of labeled cells, and it is suggested that such evidence provides a useful way of dividing the cortex into areas. By this criterion, a single, relatively large, cortical area was identified that occupied most of the medial bank of the suprasylvian sulcus, all or most of its posterior bank, and a small segment of its lateral bank. It was referred to as the Clare-Bishop area. Because neighboring visual areas were found to lack input from area 17, while the Clare-Bishop area received a strong striate input, its boundaries were investigated by labeling afferents from area 17. Together with the results of retrograde tracer injections, these data suggested that the Clare-Bishop area cuts across several of the visual areas defined physiologically by Tusa et al. ('81). As a consequence, its retinotopic organization must be relatively complex, with duplications of some parts of the visual field. Three other visual areas were tentatively identified on the basis of their distinctive connections. One was situated on the lateral bank of the suprasylvian sulcus and appeared to border the Clare-Bishop area laterally. Another, referred to as area 21, lay adjacent to area 19, and, for part of its length, also appeared to bound the Clare-Bishop area. The third, corresponding approximately to Heath and Jones's ('71) posterior suprasylvian region, lay lateral and anterior to the Clare-Bishop area in the depths of the posterior suprasylvian sulcus.

The morphology and divergent axonal organization of midbrain raphe projection neurons in the rat

The morphology of dorsal raphe neurons was examined using intracellular injections of horseradish peroxidase (HRP) and the Golgi technique. Light microscopic examination of HRP-labeled projection neurons revealed a neuron type with radiating, poorly branched and sparsely spined dendrites and terminal dendritic thickets. The stem axon of these neurons left the nucleus ventrally but gave off a beaded collateral while still within the parent cell's dendritic domain. Somatodendritic morphology from Golgi-Kopsch stained material coincided with intracellular HRP findings and the dorsal raphe may consist of varieties of one basic morphological type of neuron. Intracellular recordings made during the HRP injection experiments confirmed that stimulation of the ventral medial tegmentum elicited an antidromic action potential and an inhibitory postsynaptic potential in dorsal raphe projection neurons. The order of axonal projections arising from the midbrain raphe nuclei was examined using a double retrograde axonal tracing technique. After paired HRP and [3H] wheat germ agglutinin injections within certain projection targets of the dorsal and median raphe neurons (caudate-putamen, amygdala, hippocampus, substantia nigra and locus coeruleus), each target structure was found to have its own unique representation within a topographically distinct portion of one or more of the raphe subgroups. Neurons projecting to the caudate-putamen and substantia nigra occupied rather rostral portions. Neurons projecting to the hippocampus and locus coeruleus resided more caudally. Neurons projecting to the amygdala were situated intermediately. Overall, rostrocaudal topography in the intranuclear distributions of raphe projection neurons resulted in the formation of complex overlap zones where collateralized neurons always resided.