Retrograde Horseradish Peroxidase Method Research Articles

The neuranatomical basis of central control of cardiorespiratory interactions in vertebrates.

The neuranatomical basis of central control of cardiorespiratory interactions in vertebrates.

Cerebellovestibular projection from the posterior lobe cortex in the rabbit: an experimental study with the retrograde HRP method. I. Topographical relationships.

The sources of corticovestibular projections from the cerebellar posterior lobe vermis and hemisphere in the rabbit were investigated by the retrograde horseradish peroxidase (HRP) tracing method. Following iontophoretic injections into various subdivisions of the vestibular nuclear complex (VNC), labelled Purkinje neurones were identified ipsilaterally in all vermal lobules (VI through IX) and some regions of the hemisphere. The results indicate that projection is profuse and directed to all four nuclei of VNC. The labelling suggests some topographical relationships between lobules and especially sublobules of the posterior lobe and individual nuclei of VNC. The major projection originates from lobules VI and IX. A moderate projection derives from lobule VIII whereas that from lobule VII is small. Projections from the hemisphere arise mainly from crus I and crus II of the ansiform lobule. Those from the copula pyramidis, lobule HV and the ventral paraflocculus are sparse. In the present study a topographical relationships were more precisely established than before because the technique revealed VNC afferents from individual sublobules of the cerebellar posterior lobe cortex.

Cerebellovestibular projection from the posterior lobe cortex in the rabbit: an experimental study with the retrograde HRP method. II. Zonal organization.

The mediolateral distribution of neurones of origin of the corticovestibular projection from the cerebellar posterior lobe in the rabbit was studied with the retrograde horseradish peroxidase (HRP) tracing method. After iontophoretic injections confined to various subdivisions of the vestibular nuclear complex (VNC) labelled Purkinje neurones were found ipsilaterally in widespread mediolateral cortical regions of vermal lobules from VI to IX, as well as in crus I and crus II of the ansiform lobule, the copula pyramidis, lobule HV and the ventral paraflocculus. However, when projections from individual sublobules were examined, a clearly visible zonal pattern was found. Thus, Purkinje neurones projecting to VNC were arranged in longitudinal bands with medial and lateral boundaries quite well demarcated. These bands had a different width and were found to be located at different distances from the midline. The present findings constitute the first indication of a zonal organization of corticovestibular projections from individual sublobules of the cerebellar posterior lobe in the rabbit. This zonal arrangement of VNC afferents is discussed with the special emphasis on the cerebellar longitudinal zones and related to previous studies on other species.

The reticulovestibular projection in the rabbit: an experimental study with the retrograde horseradish peroxidase method

The reticulovestibular projections of the brainstem in the rabbit were studied by the retrograde transport of horseradish peroxidase (HRP). After selective iontophoretic injections of the tracer into various subdivisions of the vestibular nuclear complex (VNC), labeled neurons were found in defined regions of the reticular formation (RF) of the caudal pons and the rostral medulla. The results indicate that all four vestibular nuclei receive projection from RF. This projection is bilateral with a contralateral predominance. The major projection originates from dorsal and dorsolateral regions of the caudal pontine reticular nucleus (RPc) and the gigantocellular reticular nucleus (RGc) at the transitional level between them. A modest projection originates from pars α of the caudal pontine reticular nucleus (RPcα), the parvocellular reticular nucleus (Rpc) and pars α of the parvocellular nucleus (Rpcα), mostly from their ventral regions. A small projection arises from pars α of the gigantocellular reticular nucleus (RGcα), as well as from the ventral reticular subnucleus (Rv) and cell group a in the caudal aspect of the medulla. No clear-cut topical relationship was noted between the location of neurons in RF and projection site in VNC. The superior vestibular nucleus (SV) and the medial vestibular nucleus (MV) receive projections exclusively from RPc and RGc, whereas the lateral reticular nucleus (LV) and the inferior vestibular nuclues (IV) receive additional projections from the remaining RF nuclei. The termination areas of reticular fibers within SV and IV seem to be diffuse but in MV and LV there is a clear preponderance to the regions located ventrally. The present study has established cells of origin for the reticulovestibular projections from the pontomedullary RF to individual VNC nuclei in the rabbit.

Musculotopic organization of the facial nucleus of the reeler mutant mouse

The migration of facial motoneurons is affected by the reeler gene, and the facial nucleus of the reeler mutant is cytoarchitecturally abnormal. The present study was undertaken to compare the musculotopic organization of the reeler facial nucleus with that of the normal mouse by the retrograde horseradish peroxidase method. In the normal mouse, motoneurons supplying the nasolabial muscle were located in the lateral and dorsolateral subnuclei, those supplying the posterior auricular muscle in the ventromedial and dorsomedial subnuclei, those supplying the mentalis/platysma muscle in the ventral intermediate subnucleus of the facial nucleus, and those supplying the posterior belly of the digastric muscle in the accessory facial nucleus. This musculotopic representation on the main facial nucleus and accessory facial nucleus also appears in the reeler mouse. The musculotopic representation of the facial nucleus of the reeler mouse is thus identical to that of the normal mouse in spite of the former's cytoarchitectonic abnormalities.

Non-reciprocal connections between the vestibular nuclei and the cerebellar paramedian lobule, with special reference to the climbing fiber zones: an analysis in the rabbit using the retrograde horseradish peroxidase method

The organization of the secondary vestibular projections onto the cerebellar paramedian lobule (PML) and possible reciprocal corticovestibular connections were investigated by the retrograde horseradish (HRP) technique in the rabbit. Following injections of the tracer into the vestibular nuclear complex (VNC), an ill-defined, sagittal band composed of numerous labelled Purkinje cells was found ipsilaterally throughout the length of the lateral portion of the vermis, the ventral paraflocculus and the flocculus. However, no labelled Purkinje cells were found in the cortex of the PML. The results indicate that zone B, considered to give rise to cerebellar corticovestibular projections, is not present in the rabbit PML. After injections of HRP into the PML, the retrograde labelling pattern in the VNC was analyzed, in relation to the climbing fiber zones identified by retrograde labelling in the inferior olive. No clear-cut correspondence could be found between the vestibular subdivisions and climbing fiber zones in the PML, except that only the interstitial nucleus of the vestibular nerve projects into zone D 1 in sublobules e and d. There were no vestibular projections to zone D 2. The only salient feature was that cells projecting onto zones C 2, C 1 and C 3 of the PML were arranged rostrocaudally in the inferior vestibular nucleus and the caudal portion of the medial vestibular nucleus. In addition, a topical relationship was found between parts of the VNC and sublobules of the PML.

The distribution of infrahyoid motoneurons in the cat: a retrograde horseradish peroxidase study.

The distribution of cell bodies and the peripheral course of axons of infrahyoid motoneurons were examined in the cat by the retrograde horseradish peroxidase method after application of the enzyme to the peripheral nerve branches supplying the infrahyoid muscles. Infrahyoid motoneurons were observed to constitute a slender cell column, which extended from a level of the caudal part of the hypoglossal nucleus usually to the most caudal level of the C1 cord segment, or occasionally to the lower levels of the C2 cord segment. The cell column was located immediately lateral to that of motoneurons of the spinal accessory nerve. In the cell column, thyrohyoid motoneurons were distributed in the medulla oblongata; sternohyoid motoneurons were located somewhat more cranially than sternothyroid motoneurons in the medulla oblongata and cervical cord. However, the level of craniocaudal distribution of thyrohyoid, sternohyoid or sternothyroid motoneurons highly overlapped. The experiments involving severance of the hypoglossal and/or cervical nerves indicated that axons of thyrohyoid and sternohyoid motoneurons passed via the roots of both hypoglossal and C1 nerves, that axons of sternohyoid motoneurons passed via the C1 nerve roots, and that axons of infrahyoid motoneurons innervating the conjugated part of the sternohyoid and sternothyroid muscles passed usually via the C1 nerve roots, or occasionally via the roots of both C1 and C2 nerves.

Distribution pattern of lumbar epaxial, especially M. Multifidus motoneurons in the spinal cord of the cat: A study by the retrograde horseradish peroxidase method

Distribution patterns of motoneurons supplying the lumbar epaxial muscles in the spinal ventral horn have been investigated by the retrograde horseradish peroxidase (HRP) method in 6 cats. HRP (30-40%, 3.0-5.7 mg) injections into multifidus muscle in 5 cases, and for comparison, longissimus lumborum muscle in one case were made at vertebral levels from L5 to L7 at multiple sites. After two days, the animals were sacrificed and frozen sections (60 microns thick) in the frontal plane of the spinal cord were made from the L1 to L7 segments and each section reacted histochemically with the tetramethylbenzidine (TMB) method. The results are summarized as follows. Following HRP injections into the multifidus muscle, labeled cells were found in the ventral horn rostrocaudally from the L2 to L6 segments. Total number of labeled cells ranged from 176 to 436 on the injected side. Labeled cells were most numerous in the L4 segment (range: 98-244). Labeled cells were found on the medial side of the ventral horn, especially in the ventromedial part (VM), predominantly ipsilateral to the injected muscle. Following HRP injection into the longissimus lumborum muscle in one case, labeled cells were also found on the medial side, but were more restricted in the centromedial part (CM) of the ventral horn. In all cases with HRP injection into the multifidus muscle, labeled cells were occasionally observed in VM and/or the medial ventral column (VC) contralateral to the injected muscle in L3 and/or L4 segment. The total number of these cells ranged from 2 to 66. These contralaterally labeled cells were presumed to be due to the peripheral HRP spread. Labeled cells observed in VM and CM were fusiform and round in shape, respectively. These cells had mean average soma diameters (ASD) from 32 to 40 microns and were presumed to be alpha motoneurons. A few labeled cells observed in VC were fusiform in shape and were significantly smaller (mean ASD: 23-28 microns) than the cells observed in other areas (VM, CM). In some cases, the dendrites of labeled cells in VM and VC extended dorsomedially in the VC across the midline in the anterior white commissure.

Central representation of the hindlimb muscles supplied by the common peroneal nerve. A retrograde horseradish peroxidase study in the dog

Motoneurons supplying the common peroneal nerve in the dog were identified by the retrograde horseradish peroxidase method. They were distributed within two discrete cell columns (columns 2 and 2′) in the 6th and 7th lumbar segments. The extensor digitorum longus muscle was represented in the dorsolateral part of column 2; the peroneus longus muscle in the ventrolateral part and the tibialis cranialis muscle in the intermediate lateral part. The medial half of column 2 contained motoneurons supplying the superficial peroneal nerve. Column 2′, which was situated dorsomedially to column 2, contained motoneurons innervating the extensor digitorum brevis muscle.

Correlation of the main peripheral branches of the facial nerve with the cytoarchitectonic subdivisions of the facial nucleus in the guinea pig.

Correlation of the main peripheral branches of the facial nerve with morphological subdivisions of the facial nucleus was examined in the guinea pig by the retrograde horseradish peroxidase method. The facial nucleus of the guinea pig was divided cytoarchitectonically into the dorsolateral, lateral, intermediate, medio-intermediate, medial, and ventromedial divisions; the ventromedial division was further divided into the major, dorsal and lateral parts. Six main branches of the facial nerve were identified; the zygomatico-orbital, cervical, posterior auricular, anterior auricular, superior labial, and inferior labial branches. After applying HRP to the main branches of the facial nerve, the pattern of distribution of HRP-labelled neuronal cell bodies within the facial nucleus was examined: the dorsolateral division, dorsal part of the ventromedial division, major part of the ventromedial division, lateral part of the ventromedial division, or medial division contained the cell bodies of respectively the zygomatico-orbital, cervical, posterior auricular, anterior auricular, or superior labial branches, while each of the lateral, intermediate, and medio-intermediate divisions contained the cell bodies of both the superior labial and inferior labial branches.

Projection systems and terminal localization of dorsal column afferents: An autoradiographic and horseradish peroxidase study in the rat

Projection systems from the gracile nucleus and the cuneate nuclear complex to their terminal sites in the mesencephalon, diencephalon, and cerebellum were examined by means of anterograde autoradiography and retrograde horseradish peroxidase methods. Three projection systems emerge from the dorsal column nuclei, decussate via internal arcuate fibers, and form the contralateral medial lemniscus (ML). At the obex, some fibers split off the ML and course dorsolaterally, forming an ascending lateral system which fits the "lemniscal adjunct channel" (LAC) concept of Graybiel ('72). The ML continues rostrally as the "main lemniscal line channel" (MLLC). At the inferior colliculus, some LAC fibers terminate in the pontine nuclei, parabrachial, dorsal reticular nuclei, and the external and ventral medial part of the central nucleus of the inferior colliculus. More rostrally at the level of the superior colliculus, terminal fields are found in the medial nucleus of the medial geniculate body, the suprageniculate, pretectal, and mesencephalic reticular nuclei, marking the end of the LAC. In the diencephalon, gracile fibers leave the MLLC and form a crescentlike terminal field along the extreme lateral border of the ventral posterior lateral nucleus (VPL) of the thalamus. Cuneate MLLC fibers terminate in a bandlike formation in the VPL medial to the gracile termination. The third fiber system, the cuneocerebellar projection, emerges from the cuneate, the external cuneate nuclei, and the "cellular bridge" and immediately enters the ipsilateral inferior cerebellar peduncle. Upon entering the cerebellum, the major fiber component remains ipsilateral and terminates as vertical bands in vermal and paravermal lobules, and lobules I through IVa. The posterior cerebellar lobe contains terminal bands in lobules VII-IX, the copula pyramidis, and the paramedian lobule. It is concluded that the dorsolateral fiber system conforms to Graybiel's LAC. It is more divergent and probably less modality specific, whereas the medial lemniscal system conforms to the MLLC, which is said to be modality specific, less divergent, and locked to specific sensory-motor response characteristics. The topography of cerebellar terminal bands indicates that there is sensory-motor representation from all parts of the body to all parts of the cerebellum, at least in the rat.

Cells of origin and topographic organization of corticospinal neurons in the guinea pig by the retrograde HRP method

In the guinea pig, horseradish peroxidase (HRP) was injected in the cervical or lumbar enlargements of the spinal cord in order to examine the origin and topographical organization of corticospinal (CS) neurons. The cortex was divided into granular and agranular regions to attempt correlations with the location of labeled CS neurons. These, of all sizes, are found only in pyramidal layer V of both kinds of cortical regions and could be seen as single, grouped or organized in clusters of 3–5 or more cells. The soma diameters of HRP CS neurons ranged from 13 to 49 μm. The largest labeled cells were present in the medial part of both agranular and granular cortices and included the giant pyramidal neurons which were found only in the rostromedial agranular cortex. The predominating intermediate-size cells were mostly present in lateral granular areas. The smaller cells were distributed in rostrolateral agranular and caudal granular regions. Morphological evidence suggests that an aggregation in clusters of labeled neurons and the different groups of CS neurons identified in different cortical areas may have additional differences in organization with respect to their precise topographical relations and functional properties. Neurons projecting to cervical levels were more abundant compared to projecting to the lumbar spinal cord, being distributed widely more lateral and anterior on the hemisphere to cortical neurons projecting to lumbar enlargement. These were located only medially on the dorsal surface of the brain. The two groups of labeled neurons were distributed in both agranular and granular cortices and occur adjacent to each other with only a narrow strip of overlap. The findings i.e. laminar organization, the pyramidal shape of CS neurons of all sizes, the presence of clusters and the somatotopic distribution of CS neurons seen in the guinea pig are discussed in comparison with equivalent findings on the corticospinal tract (CST) system of other species. The presence of labeled neurons within the cortex on both sides following unilateral HRP injections in the spinal cord points to a bilateral origin of the CST. This findings is discussed on the basis of the present study and previous autoradiographic findings.

Topographical projections from the thalamus to the putamen in the cat

Thalamic projections to the putamen (Put) in the cat were studied by the retrograde horseradish peroxidase method. Major thalamic projections to the Put originate from the midline and intralaminar nuclear regions including the centre médian-parafascicular complex (CM-Pf). The other thalamic projections to the Put arise mainly from the suprageniculate nucleus (Sg), magnocellular division of the medial geniculate nucleus (MGm), caudomedial part of the lateroposterior nucleus (LP) and ventrolateral part of the ventromedial nucleus (VM). The VM projects to the rostral Put, while the posterior thalamic regions (Sg, MGm, LP) project to the caudal Put.

The localization of vagal neurons in the terrapin ( Trionyx sinensis) as revealed by the retrograde horseradish peroxidase method

After the application of horseradish peroxidase in Nonidet P40 solution to the right cervical vagus in the terrapin, the neurons giving origin to efferent fibers and the transganglionic sensory fibres are labeled. The neurons forming the vagus nerve at the mid-cervical level are located in the dorsal motor nucleus of the vagus nerve (DMX), in the nucleus ambiguus (NA), in an area located ventral to the DMX, and less definitely in the nucleus of the spinal accessory nerve. The DMX and NA neurons are bilaterally distributed with a predominant ipsilateral component. Labeled DMX neurons appear to have a slightly more extensive distribution than those in mammals and their dendrites extend to the nucleus of the tractus solitarius, hypoglossal nucleus (HN), lateral wall and floor of the 4th ventricle and a region dorsomedial to the HN. Some recurrent collaterals of the DMX neurons loop round the medial longitudinal fasciculus to end in a region immediately ventrolateral to the DMX. Transganglionic sensory fibers course with the tractus solitarius and spinal tract of the trigeminal nerve to end in the respective nucleus associated with the tract.

Direct projections from the substantia nigra to the posterior thalamic regions in the cat

The substantia nigra (SN) of the cat was shown, by the anterograde and retrograde horseradish peroxidase methods, to contain neurons which send their axons to the caudomedial portions of the suprageniculate nucleus and/or lateroposterior nucleus of the thalamus; these neurons were located in the restricted region in the lateral part of the SN, which corresponds to the pars lateralis of the SN.

Observations on the brainstem-spinal descending systems of normal and reeler mutant mice by the retrograde HRP method.

Brainstem neurons which project to the lumbar spinal level were identified in both reeler mutant mice and normal controls (Balb/c mice) by the retrograde horseradish peroxidase (HRP) technique. In normal controls after HRP injection into the lumbar cord, retrogradely labelled neurons were observed in (1) the lateral vestibular nucleus, (2) the pontine and medullary reticular formations including the nucleus centralis caudalis pontis, nucleus gigantocellularis, nucleus paragigantocellularis, nucleus raphe magnus et pallidus, and nucleus centralis medullae oblongatae pars ventralis et dorsalis, and (3) the dorsal column nuclei, i.e., the nucleus gracilis and nucleus cuneatus medialis. In reeler mutant mice, labelled neurons were again seen in the nuclei referred to above, and their cellular type and distribution patterns within the corresponding nuclei were similar to those of the normal controls. These observations suggest that (1) the brainstem nuclei of reeler mutant mice which project to the lumbar spinal cord are cytoarchitecturally normal, (2) the reeler genetic locus (rl) does not affect the nonlaminated structures in the brainstem, at least those referred to above, and (3) the motor dysfunctions observed in the reeler, such as action tremor, dystonic posture, and reeling ataxic gait, are not attributable to the brainstem-spinal descending systems.

Direct projections to the prestriate cortex from the retino-recipient zone of the inferior pulvinar nucleus in the macaque monkey

It was found by the anterograde and retrograde horseradish peroxidase methods that the medial border region of the inferior pulvinar nucleus of the Japanese monkey ( Macaca fuscata), where optic fibers have been shown to end [8], was connected reciprocally with the ipsilateral prestriate cortical regions around the compensatory sulcus on the medial surface of the cerebral hemisphere.

The cells of origin of the incertofugal projections to the tectum, thalamus, tegmentum and spinal cord in the rat: A study using the autoradiographic and horseradish peroxidase methods

Efferent projections of the zona incerta were examined in the rat using the autoradiographic and horseradish peroxidase methods, with special reference to the cytoarchitectonic structure of the zona incerta. Autoradiographic experiments showed that the incertofugal fiber systems reach ipsilaterally to the thalamus (lateral dorsal, central lateral, ventral lateral geniculate, parafascicular, subparafascicular and reuniens nuclei, and posterior nuclear complex), to the hypothalamus (dorsal, lateral and posterior hypothalamic areas), to the tectum (medial pretectal area, deep pretectal and pretectal nuclei, superior colliculus and periaqueductal gray) and to the midbrain tegmentum, pons and medulla oblongata (subcuneiform, cuneiform and red nuclei, nuclei of the posterior commissure and Darkschewitsch, interstitial nucleus of Cajal, pedunculopontine tegmental nucleus, oral and caudal pontine reticular nuclei, nucleus raphe magnus, gigantocellular reticular nucleus, pontine gray and inferior olivary complex). Contralaterally, incertal efferent fibers reach to the zona incerta. Cells of origin of the incertofugal fiber systems to the tectum, thalamus, tegmentum and spinal cord were examined using the retrograde horseradish peroxidase method. Cells of origin of the incertotectal pathway are located mainly in the ventral and caudal parts of the zona incerta and partly in the antero-polar, dorsal and postero-polar parts. Cells projecting to the thalamus (at least to the lateral dorsal and central lateral nuclei) are situated in the ventral and caudal parts of the zona incerta, but they are rare in the other incertal structures. Cells of origin of the incertotegmental system are located mainly in the dorsal, magnocellular and caudal parts and partly in the antero- and postero-polar parts, but they are not situated in the ventral part. Cells of the magnocellular part project more caudally to the medulla oblongata and spinal cord than those of the other parts of the zona incerta. Forel's field contains many cells projecting to the tegmentum. The results provide good evidence that the cells of origin of efferent projections are topographically organized and are related to cytoarchitectonic areas within the zona incerta.

Isthmic afferent neurons identified by the retrograde HRP method in a teleost, Navodon modestus

Isthmic afferent neurons were investigated by the retrograde horseradish peroxidase (HRP) method in a teleost, Navodon modestus. Following HRP injections into the nucleus isthmi, large pyriform neurons are labeled in the ipsilateral optic tectum. Very large and multipolar neurons are also labeled in the ipsilateral nucleus pretectalis. No labeled neurons were found in other areas.

The density, distribution and topographical organization of spinocervical tract neurones in the cat.

1. In acute experiments, detailed grids of micro-electrode recordings were made from spinocervical tract (s.c.t.) cells in the lumbosacral cord of anaesthetized cats. These grids provided electrophysiological data on the location, distribution, density and somatotopic organization of s.c.t. neurones.2. In acute experiments lasting up to 48 hr, retrograde labelling of s.c.t. cells was carried out by injecting horseradish peroxidase into the lateral cervical nucleus in anaesthetized cats. The aim was to flood the nucleus with the enzyme so that all s.c.t. neurones would be labelled in order to provide an independent check on the location, density and distribution data obtained in the electrophysiological experiments.3. The electrophysiological and the anatomical experiments were sometimes performed on the same animal.4. The electrophysiological and anatomical results were in excellent agreement. (a) S.c.t. cells are located mainly in ipsilateral laminae III, IV and V. About 25% are in lamina III, 60% in lamina IV and 10% in lamina V. There are a few cells in laminae I, II and VI making up the remaining 5%. (b) There are about 550-800 s.c.t. cells in the lumbosacral enlargement (L4-S2 inclusive) on each side of the cord. Most cells are in L7-S1 where there are twenty to forty s.c.t. neurones in each millimetre length of cord.5. Many marginal (lamina I) cells were labelled with the retrograde horseradish peroxidase method and a few cells on the contralateral side in laminae III-V and VII-VIII were also labelled. The marginal cells formed 12-18% of labelled cells on the side of injection and, in addition, a similar absolute number of marginal cells was labelled on the side contralateral to the injection. The possible identity of these neurones is discussed.6. S.c.t. cells form a sheet of neurones across the dorsal horn. The sheet is organized somatotopically in a way which resembles the representation of the hind limb dermatomes in the dorsal columns (Werner & Whitsel, 1967). There is a relative enlargement of the L6-S1 dermatomes which encroach upon their neighbouring cord segments. The gradient of the map is very steep in the mediolateral direction but gradual in the rostrocaudal direction. The detailed somatotopic maps generated in the present work have revealed that s.c.t. cells are arranged so that their receptive fields form longitudinal columns and cells within the columns have overlapping fields.