Postsynaptic dorsal column neurons in the cat: a study with retrograde transport of horseradish peroxidase

Summary In situ hybridization histochemistry and RNA blots were used to study the expression of glutamic acid decarboxylase (GAD) mRNA in rats with or without a unilateral lesion of midbrain dopamine neurons. Two populations of GAD mRNA positive neurons were found in the intact caudate-putamen, substantia nigra and fronto-parietal cortex. In caudate-putamen, only one out of ten of the GAD mRNA positive neurons expressed high levels, while in substantia nigra every second of the positive neurons expressed high levels of GAD mRNA. Relatively few, but intensively labelled neurons were found in the intact fronto-parietal cerebral cortex. In addition, one out of six of the GAD mRNA positive neurons in the fronto-parietal cortex showed a low labeling. On the ipsilateral side, the forebrain dopamine deafferentation induced an increase in the number of neurons expressing high levels of GAD mRNA in caudateputamen, and a decrease in fronto-parietal cortex. A smaller decrease was also seen in substantia nigra. However, the total number of GAD mRNA positive neurons were not significantly changed in any of these brain regions. The changes in the levels of GAD mRNA after the dopamine lesion were confirmed by RNA blot analysis. Hence, midbrain dopamine neurons appear to control neuronal expression of GAD mRNA by a tonic down-regulation in a fraction of GAD mRNA positive neurons in caudate-putamen, and a tonic up-regulation in a fraction of GAD mRNA positive neurons in fronto-parietal cortex and substantia nigra.     

Electron microscopic observations on the synaptology of cat sciatic gamma-motoneurons after intracellular staining with horseradish peroxidase

Four cat sciatic motoneurons with axon conduction velocities below 30 m/s, and thus considered to be of the gamma-type, were intracellularly labelled with horseradish peroxidase (HRP) and subsequently studied in the electron microscope. The labelled neurons were apposed by synaptic terminals with spherical (S-type) and flattened vesicles (F-type) but not by large terminals with spherical vesicles (M- and C-types) seen on alpha-motoneurons. Quantitative analysis of a complete series of ultrathin sections through one of the neurons showed that the synaptic covering on the cell body (24.2%) was considerably larger than what has been reported for triceps surae gamma-motoneurons, but within the range of values for gamma-motoneurons in the thoracic region of the spinal cord.     

The septohippocampal projection in the rat: an electron microscopic horseradish peroxidase study

In order to identify the specific targets of the septohippocampal projection in the rat, horseradish peroxidase localization at the electron microscopic level was used. Following injections of free horseradish peroxidase into the medial septum, sections of the dorsal hippocampal formation were reacted with diaminobenzidine and processed for electron microscopy by routine methods. Sections were viewed unstained. Horseradish peroxidase labeling in the dentate gyrus was predominantly in the supra- and infragranular layers. All postsynaptic elements were neuronal. They included granule cell somata and somata and dendrites of hilar cells; these may include pyramidal basket cells. No synaptic contacts with vascular or glial elements were found. These results provide a basis for comparing the specific targets of the septohippocampal projection with those of the sympathohippocampal pathway, which innervates the dentate following lesions of the septohippocampal projection.     

Spinocervical tract cells in cat and dog,labeled by the retrograde transport of horseradish peroxidase

The location of cells of origin of the spinocervical tract has been determined in cat and dog by injecting horseradish peroxidase into the lateral cervical nucleus (LCN). Retrogradely labeled cells were found predominantly in the ipsilateral nucleus proprius of the dorsal horn along the entire spinal cord. Cells of Clarke's column and contralateral ventral horn cells were not labeled, in agreement with previous electrophysiological indications that the spinocerebellar tracts do not maintain collateral terminations in the LCN.     

The trigemino-olivary projection in the cat as studied with retrograde transport of horseradish peroxidase

Summary The trigemino-olivary projection was studied in cats which from a ventral approach were injected with horseradish peroxidase into various parts of the inferior olive. Retrogradely labelled cells of all sizes were present in all three divisions of the spinal sensory nucleus: nucleus caudalis, nucleus interpolaris and nucleus oralis. The projection is bilateral with the highest number of labelled cells on the contralateral side. No retrogradely labelled cells are found in the principal nucleus, but some cats showed a few retrogradely labelled cells within the ipsilateral mesencephalic nucleus.The findings are discussed and related to recent observations concerning the distribution of the direct trigemino-cerebellar fibres.     

The reticulocerebellar projection in the cat as studied with retrograde transport of horseradish peroxidase

Summary Injections of horseradish peroxidase into the various parts of the cerebellar cortex and the cerebellar nuclei in the cat result in labelled cells within the reticular formation proper. All the reticular nuclei (with the exception of the reticular formation of the mesencephalon) send fibres to the cerebellum. The highest number of labelled neurons after cerebellar injections is found in the caudal reticular formation, especially within nucleus reticularis ventralis, nucleus reticularis lateralis and nucleus reticularis gigantocellularis. Another region for an accumulation of labelled cells is the rostral part of nucleus reticularis pontis caudalis.Except for the paraflocculus, all cerebellar cortical areas and all cerebellar nuclei receive afferents from one or more of the nuclei within the reticular formation proper, but the largest number of labelled neurons is observed in cases with injections including the intermediate-lateral part of lobulus simplex and the adjacent areas of the anterior lobe and crus I. The projection is bilateral with an ipsilateral preponderance (the cerebellar nuclei appear to receive a higher number of fibres from the contralateral side). Cells of all sizes are labelled, but labelled giant cells are found only after large cortical injections.     

The diencephalo-olivary projection in the cat as studied with retrograde transport of horseradish peroxidase

Summary The projection from certain diencephalic regions (zona incerta, Forel's fields, the parafascicular and subparafascicular nuclei, the periventricular grey and the hypothalamus) to the inferior olive in the cat was studied by means of retrograde protein tracing. Small injections of horseradish peroxidase were made into various parts of the inferior olive from a ventral approach. The number of retrogradely labelled neural cells in the diencephalic nuclei of all cats is presented in Table 2. The majority of the labelled cells was found in the parafascicular and subparafascicular nuclei, especially within the medial part of the former. The connection is ipsilateral, and the caudal as well as the rostral part of the olivary complex appears to receive the descending afferents. The findings are discussed and related to recent observations concerning descending afferents to the olivary complex.     

The vestibulothalamic projections in the cat studied by retrograde axonal transport of horseradish peroxidase

Summary Horseradish peroxidase (HRP) was injected or iontophoretically ejected in various thalamic nuclei in 63 adult cats. In 11 other animals HRP was deposited outside the thalamic territory. The number and distribution of labelled cells within the vestibular nuclear complex (VC) were mapped in each case. To a varying degree all subgroups of VC appear to contribute to the vestibulothalamic projections. Such fibres are distributed to several thalamic areas. From the present investigation it appears that generally speaking, there exist three distinct vestibulothalamic pathways with regard to origin as well as to site of termination of the fibres. One projection appears to originate mainly in caudal parts of the medial (M) and descending (D) vestibular nuclei and in cell group z. This pathway terminates chiefly in the contralateral medial part of the posterior nucleus of the thalamus (POm) including the magnocellular part of the medial geniculate body (Mgmc), the ventrobasal complex (VB) and the area of the ventral lateral nucleus (VL) bordering on VB. A second projection originates mainly in the superior vestibular nucleus (S) and in cell group y and terminates mainly in the contralateral nucleus centralis lateralis (CL) and the adjoining nucleus paracentralis (Pc). A third, more modest, pathway originates chiefly in the middle M and D, with a minor contribution from S and cell group y, and terminates in the contralateral ventral nucleus of the lateral geniculate body (GLV). There is some degree of overlap between the origin of these three vestibulothalamic pathways.Abbreviations B.c. brachium conjunctivum - CeM nucleus centralis medialis thalami - CL nucleus centralis lateralis thalami - CM nucleus centrum medianum - D nucleus vestibularis descendons - f cell group f - g cell group g - GLD corpus geniculatum laterale dorsalis - GLV corpus geniculatum laterale ventralis - i.e. nucleus intercalatus - L nucleus vestibularis lateralis - LD nucleus lateralis dorsalis thalami - LIM lamina medullaris interna - Lim nucleus limitans - LP nucleus lateralis posterior thalami - M nucleus vestibularis medialis - MD nucleus medialis dorsalis thalami - MGmc corpus geniculatum mediale, pars magnocellularis - MGp corpus geniculatum mediale, pars principalis - N.cu.e. nucleus cuneatus externus - N.f.c. nucleus fasciculi cuneati - N.mes. V nucleus mesencephalicus nervi trigemini - NR nucleus ruber - N.tr.s. nucleus tractus solitarius - N. VII nervus facialis - N. VIII nervus statoacusticus - PC pedunculus cerebri - Pc nucleus paracentralis thalami - Pf nucleus parafascicularis - p.h. nucleus prepositus hypoglossi - PO posterior thalamic group - PO1 lateral part of PO - POm medial part of PO - Prt nucleus pretectalis - Pul pulvinar - R nucleus reticularis thalami - S nucleus vestibularis superior - Sg nucleus suprageniculatus - SN substantia nigra - Sv nucleus supravestibularis - Tr.s. tractus solitarius - VA nucleus ventralis anterior thalami - VL nucleus ventralis lateralis thalami - VPL nucleus ventralis posterior lateralis - VPL1 lateral part of VPL - VPLm medial part of VPL - VPM nucleus ventralis posterior medialis - x cell group x - y cell group y - z cell group z - V nucleus motorius nerve trigemini - X nucleus dorsalis nerve vagi - XII nucleus nervi hypoglossi     

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

Summary 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.     

Pallido-cortical projections in the cat studied by means of the horseradish peroxidase retrograde transport technique

Using a multi-microelectrode, in 5 animals, orientation tuning was measured simultaneously in 30 closely spaced parallel penetrations perpendicular to the surface of the striate cortex. Actual penetration angles were determined by three-dimensional track reconstruction. Above and below layer IVc, two columnar systems were found whose orientation angles were independent.     

Cortical projections to the periaqueductal grey in the cat: a retrograde horseradish peroxidase study

Stereotaxic fluid microinjections of horseradish peroxidase into different parts of the rostral and caudal periaqueductal grey (PAG) in cats have provided substantial retrograde evidence that the somatosensory cortex (I and II), frontal cortex, insular and cingular cortex are the principal sources of cortical-PAG projections. The somatosensory cortex II projects to all the regions of the rostral and caudal PAG. The frontal cortex projects to dorso-lateral quadrant of the PAG. The same projections were determined from insular and cingular cortex to PAG. The findings revealed a morphological substratum of corticofugal effects on PAG.     

Ultrastructure of visual callosal neurons in cat identified by retrograde axonal transport of horseradish peroxidase

Summary The ultrastructure of neurons at the border of areas 17 and 18 of the visual cortex of the cat was studied by the combined use of the retrograde transport of horseradish peroxidase (HRP) and electron microscopy. Callosal neurons were retrogradely labelled by injecting HRP at the 17/18 border region of the contralateral hemisphere. They were found mainly in layer III but also in IV and VI. They were most commonly pyramidal cells and less often large, spiny stellate cells. Pyramidal callosal neurons received only symmetrical synapses on their soma and mainly symmetrical (but a few asymmetrical) synapses on their dendritic shafts. Their abundant spines received asymmetrical synapses. The stellate cells were contacted by moderate numbers of symmetrical and asymmetrical axodendritic and axosomatic synapses and also had asymmetrical axospinous contacts. We propose that the callosal stellate neurons consist of a class of large spiny stellates, recognizable by light and electron microscopic criteria.The work described in this paper forms part of a study for a doctoral dissertation in the University of Lausanne by J. P. Hornung.     

The efferent vestibular system in the cat: a horseradish peroxidase and fluorescent retrograde tracers study

Neurons of the efferent vestibular system were investigated in the cat using retrograde axonal transport of horseradish peroxidase and fluorescent retrograde double labelling techniques. The number of efferent neurons was clearly higher than previously reported. A three dimensional reconstruction of the location of these neurons showed that they constitute a single group and did not give evidence of an eventual specialization based on neuron subpopulations. However, a study of cross-sectional areas of the horseradish peroxidase-labelled efferent neurons detected that the ipsilateral population contained a larger number of small neurons than the contralateral one. Double labelling by means of either 4',6-diamidino-2- phenylindol 2HCl in combination with horseradish peroxidase or Fast Blue in combination with Nuclear Yellow showed that 20% of efferent neurons project to both labyrinths. Such a high percentage raises the question of the role of these double-projecting cells and the specificity of their branching on vestibular receptors. This study expands previous work in the cat demonstrating that a much greater number of efferent neurons exists than had hitherto been assumed, among them 20% have both crossed and uncrossed projections.     

Identification of axon terminals of the cerebello-olivary fibers in the cat: an electron microscope study using the anterograde horseradish peroxidase method

After injection of horseradish peroxidase (HRP) into the lateral cerebellar nucleus of the cat, axon terminals containing electron-dense HRP-granules were identified contralaterally within the principal olive. These terminals contained spherical synaptic vesicles, and made asymmetrical synaptic contacts with dendritic profiles of small or medium size.     

The afferent connections of the inferior olivary complex in rats: A study using the retrograde transport of horseradish peroxidase

Retrograde transport of horseradish peroxidase (HRP) was used to define the origin of afferents to the inferior olivary complex (IOC) in rats. Using both ventral and dorsal surgical approaches to the brainstem, HRP was injected into the IOC through a micropipette affixed to the tip of a 1-μl Hamilton syringe. After a 2-day postoperative survival, animals were sacrificed by transcardiac perfusion with a 1% paraformaldehyde-1.25% gluteraldehyde solution, and brains were processed according to the DeOlmos protocol (1977), using o-dianisidine as the chromogen. Labeled cells were found at many levels of the nervous system extending from lumbar spinal cord to cerebral cortex. This wide-ranging input from numerous regions clearly underscores the complexity of the IOC and its apparent involvement in several functions. Within the spinal cord, labeled neurons were identified from cervical to lumbar but not at sacral levels. These neurons were found contralaterally in the neck region of the dorsal horn and in the medial portions of the intermediate gray. In the caudal brainstem, reactive cells in the dorsal column nuclei, the spinal trigeminal nucleus, and the subnucleus y of the vestibular complex were observed primarily contralateral to the injection sites. Labeling within the gigantocellular, magnocellular, ventral, and lateral reticular nuclei and the nucleus prepositus hypoglossi was primarily ipsilateral. Reactive neurons in the medial and inferior vestibular nuclei were predominantly ipsilateral or contralateral to HRP injections into the caudal or rostral IOC, respectively. The dentate and interposed nuclei of the cerebellum contained small, lightly labeled neurons primarily contralateral to the injection site, while the fastigial nuclei contained a few relatively large, heavily labeled cells bilateral to caudal olivary injections. Ipsilaterally labeled mesencephalic regions included the periaqueductal gray, interstitial nucleus of Cajal, rostromedial red nucleus, ventral tegmental area, medial terminal nucleus of the accessory optic tract, nucleus of the optic tract, and the lateral deep mesencephalic nucleus. The caudal part of the pretectum and small cells of the stratum profundum of the superior colliculus were labeled predominantly contralateral to the injection. In the caudal diencephalon labeled neurons were most numerous within the nucleus of Darkschewitsch and the subparafascicular nucleus, primarily ipsilateral to olivary injections. Scattered reactive neurons were also found within the ipsilateral zone incerta. With the exception of the zona incerta, all labeled mesencephalic and diencephalic nuclei had some bilateral representation of labeled cells. No labeled neurons were identified within the basal ganglia, while numerous reactive cells were found bilaterally within layer V of the frontal and parietal cerebral cortex.     

The pontocerebellar projection in the rhesus monkey: an experimental study with retrograde axonal transport of horseradish peroxidase.

The pontocerebellar projection has been studied in the Rhesus monkey by use of the retrograde axonal transport of horseradish peroxidase. The parts of the cerebellum investigated receive afferents from pontine cell groups arranged as rostro-caudally oriented lamella or slab-like regions. As a rule labelled cells are found at all rostro-caudal levels in discrete groups, but their number at various levels, and particularly their distribution in the transverse plane, differ according to which part of the cerebellum has been injected. Although the same cell group may apparently be labelled after injections of different parts of the cerebellum, each cerebellar region has its own characteristic territory in the pontine nuclei. The anterior lobe receives fibres from lamella-like regions mainly in the caudal half of the pons, with cells projecting to the vermis and intermediate parts of the anterior lobe somewhat differently situated. Crus I is connected mainly with a region rostromedially in the pons, while crus II receives fibres from all levels of the pons, the cells being located medially and ventrally at caudal levels and more laterally at rostral levels. The paramedian lobule is supplied from pontine cells restricted to the rostral two-thirds of the pons and located more laterally than those projecting to crus II. Lobules VII and VIIIA (the main part of the vermal visual area) receive fibres mainly from two long column-like regions located dorsomedially and dorsolaterally, while lobulus VIIIB is supplied mainly from other areas in the pons.Counting of labelled cells indicates that about 30% of the fibres to the vermis, and about 10% to the hemispheres, are uncrossed. The density of the projection to the cerebellar hemispheres is found to be about three times as high as to the vermis. In two cases injections of horseradish peroxidase were placed in the cerebral cortex as well as in the cerebellum, enabling a direct comparison of sites of termination of corticopontine fibres (orthograde transport of horseradish peroxidase) with location of pontocerebellar neurons.It is suggested that the cortico-ponto-cerebellar pathway is organized so as to bring about convergence in the cerebellar cortex from several parts of the cerebral cortex, and that this takes place in a highly organized manner so that each small part of the cerebellar cortex has its own characteristic set of inputs.     

The cerebellar projection from locus coeruleus as studied with retrograde transport of horseradish peroxidase in the cat

Summary The cerebellar afferent projection from locus coeruleus has been studied in the cat by means of retrograde axonal transport of horseradish peroxidase. Labelled cells are present bilaterally in locus coeruleus only following injections in the cerebellar vermis (especially its anterior and posterior parts), the ventral paraflocculus and the flocculus. The labelled cells are restricted to the caudal half of the nucleus.A few labelled cells are also present in locus coeruleus following injections in the fastigial nucleus, and in nucleus interpositus anterior. The findings are discussed in relation to other studies on the efferent and afferent connections of the locus coeruleus.On leave from the Laboratory of Neurobiology and Department of Anatomy, Faculty of Science, Mahidol University, Bangkok, Thailand, under the Fellowship Program of the Norwegian Agency for International Development (NORAD)     

The rubro-olivary tract in the cat, as demonstrated with the method of retrograde transport of horseradish peroxidase

The rubro-olivary projection in the cat was investigated by means of the retrograde transport of horseradish peroxidase. After injections in the inferior olive, more than a thousand labelled neurons were found in the ipsilateral red nucleus. These neurons had triangular-shaped cell bodies with an average diameter of 26.4 +/- 7.7 microns (mean +/- S.E.M.) and had few dendrites. Between 85% and 95% of the rubro-olivary neurons were found in the rostral third of the red nucleus (between A 5.5 and A 7). Morphologically, the rubro-olivary neurons are similar to rubro-thalamic neurons. Previous studies with retrograde transport of horseradish peroxidase have failed to demonstrate an extensive projection from the red nucleus to the inferior olive. These results are discussed in relation to our own findings.