Horseradish Peroxidase Method Research Articles

Cerebral cortical projections to the reticular regions around the trigeminal motor nucleus in the cat.

Cerebral cortical regions which send projection fibers to the reticular regions around the trigeminal motor nucleus were identified in the cat by the horseradish peroxidase (HRP) method. The reticular region around the trigeminal motor nucleus are known to contain many interneurons for masticatory motoneurons. After injections of HRP into the reticular regions around the trigeminal motor nucleus, HRP-labeled neuronal cell bodies in the cerebral cortex were found in layer V. They were distributed bilaterally in the orbitofrontal cortical regions, mainly in the rostral extension of the orbital gyrus close to the presylvian sulcus; more were located in the floor and lateral bank of the presylvian sulcus than in the crown of the orbital gyrus. After injections of HRP conjugated with wheat germ agglutinin (WGA-HRP) into these cortical regions, many labeled presumed axon terminals were distributed bilaterally in the reticular regions around the trigeminal motor nucleus; mainly in the region ventral to the trigeminal motor nucleus and in the intertrigeminal region between the main sensory trigeminal nucleus and the trigeminal motor nucleus. Terminal labeling in these regions was more prominent after WGA-HRP injection into the lateral bank of the presylvian sulcus than after WGA-HRP injection into the crown of the orbital gyrus. Thus, the present results indicate that the main part of the cortical region projecting directly to the reticular regions around the trigeminal motor nucleus in the cat is folded into the presylvian sulcus.

Thalamic projections to motor, prefrontal, and somatosensory cortex in the sheep studied by means of the horseradish peroxidase retrograde transport method.

In this study the motor, prefrontal, and somatosensory areas of the sheep cerebral cortex were defined on the basis of their thalamic afferents traced with the horseradish peroxidase method. The motor area (areas 4 and 6) occupies the cruciate gyrus. It receives a substantial projection from the thalamic nuclei ventralis anterior, ventralis lateralis, medialis dorsalis, and centralis lateralis and a smaller one from the nuclei ventralis medialis, centralis medialis, paracentralis, lateralis dorsalis, lateralis posterior, centromedianus, parafascicularis, suprageniculatus, ventralis posterolateralis, and the midline nuclei. Area 4 receives afferents mainly from the nuclei ventralis anterior, ventralis lateralis, medialis dorsalis, and lateralis posterior, whereas area 6 receives afferents mainly from the nuclei ventralis anterior, medialis dorsalis, and lateralis posterior and fewer afferents from the nucleus ventralis medialis. The prefrontal area occupies the gyrus proreus and receives numerous afferents from the nucleus medialis dorsalis and fewer from the nuclei lateralis posterior and ventralis medialis. The area extending between the lateral fissure, the coronal sulcus, the presylvian sulcus, and the rostral branch of the lateral fissure is connected mainly with sensory thalamic nuclei. Thalamic afferents were found to emanate from the nuclei ventralis posteromedialis (its parvicellular part included), ventralis posterolateralis, ventralis medialis, paracentralis, lateralis posterior, medialis dorsalis, centromedianus, suprageniculatus, paraventricularis, the substantia nigra, and the ventral part of the lateral geniculate nucleus. The first somatosensory area (Johnson et al., '74, J. Comp. Neurol. 158:81-108) was found to extend between the coronal, the diagonal, and the anterior suprasylvian sulci and to receive afferents almost exclusively from the nucleus ventralis posteromedialis.

Origin of interhemispheric fibers in acallosal opossum (with a comparison to callosal origins in rat)

The neocortical origins of the anterior commissure in the acallosal, marsupial opossum were studied with the horseradish peroxidase (HRP) method. Following complete surgical transection of the anterior commissure, HRP was applied directly to the cut fiber tips. This procedure resulted in very large numbers of vividly labeled cells within the neocortex. The labeled cells were plotted and counted for comparison among cytoarchitectonic areas and among cortical layers. For comparative purposes, the neocortical origins of the corpus callosum are studied with the same procedure in the rat. No cytoarchitectonic area was entirely devoid of labeled cells in either species. The concentration of labeled cells throughout the entire neocortex averaged 25.2 cells/0.05 mm3 in opossum and 31.2 cells/0.05 mm3 in rat. The concentrations of labeled cells were correlated for the eight cytoarchitectonic areas common to the two species, though they were different enough in number to be statistically reliable. The distribution of labeled cells both among and within cytoarchitectonic areas was often more homogeneous in opossum than in rat. Although cortical layer 1 had no labeled cells in either species, the distribution of labeled cells across the remaining cortical layers differed sharply between the two species. In opossum, layer 3 had the most labeled cells (averaging 55% of the total number) while layer 5 had considerably less (averaging 12%). In rat, layer 5 had as many labeled cells as layer 3--both layers averaging 43% of the total number of labeled cells. In both species, striate cortex deviated markedly from other cytoarchitectonic areas. Although both species had very few labeled cells in striate cortex, those that were labeled were invariably supragranular in opossum and infragranular in rat. The similarities and dissimilarities in the topographic distribution of the origins of the two types of interhemispheric fiber systems seem to parallel the degree of cortical (and thalamic) differentiation in the two animals. However, the differences in laminar distribution are much greater and in particular, the small contribution of layer 5 in opossum as opposed to rat may well be functionally significant.

Significance of Con A binding sites in the conjugation of Euplotes vannus

AbstractAggregations of Con A binding sites that occur on the surface of preconjugant cells have been studied in order to reveal their possible functions in the conjugation process. By means of a newly adapted horseradish peroxidase method, they can be visualized in distinctly shaped oval fields (OF) as soon as 10–15 min after mixing cells of opposite mating types. Cycloheximide (CHX) prevents establishment of new OFs but does not effect preexisting ones. In the absence of previously developed OFs, cells cannot form pairs. However, even if OFs are fully present, pairing still can be inhibited by CHX, high K+ concentrations (≥ 60 mmol/liter), or the K+‐channel blocker tetraethyl‐ammonium (TEA, 10 mmol/liter). A doublet cell with OFs on both halves normally binds to one singlet. This partner, after a stay of 10 min in an angled agglutination position, quickly rotates into the parallel pairing position. At this moment, the doublet suddenly loses its affinity for complementary cells at its free OF. This behavior explains why many doublets pair only with one singlet. The Con A binding sites themselves seem not to be active in cell pairing; they are regarded as a marker for a region of the cell surface whose contact with a complementary cell subsequently results in the unmasking of unidentified compounds that serve as adhesive agents and/or the hinge for the quick rotation of agglutinated cells.

Localization of the spinal accessory motoneurons in the cervical cord in connection with the phrenic nucleus: an HRP study in cats

The localization of the spinal accessory motoneurons (SAMNs) that innervate the accessory respiratory muscles, the sternocleido-mastoid (SCM) and trapezius (TP) muscles, was identified in the cat using the horseradish peroxidase (HRP) method. In the cases of HRP bathing of the transected spinal accessory nerve (SAN), HRP-labeled motoneurons were observed ipsilaterally from the C1 to the rostral C6 segments of the spinal cord. Labeled neurons were located principally in the medial and central regions of the dorsomedial cell column of the ventral horn in the C1 segment, in the lateral region of the ventrolateral cell column in the C2–C4 segments, between the ventrolateral and ventromedial cell columns in the C5 segment and in the lateral region of the ventromedial cell column in the C6 segment. In the cases of HRP injection into either SCM or TP muscles, labeled SCM motoneurons were found in the C1–C3 segments of the spinal cord and labeled TP motoneurons were chiefly localized more caudally within the spinal accessory nucleus. The present study revealed that, in the C5 and C6 segments, the SAMNs have a very similar topographic localization to the phrenic nucleus in the ventral horn. This finding implicated the functional linkage of the SAMNs with the phrenic motoneurons in particular types of respiration.

The vestibuloocular reflex of the adult flatfish. II. Vestibulooculomotor connectivity.

The peripheral and central oculomotor organization of the adult flatfish presents no morphological substrates that suffice to explain adaptive changes in its vestibuloocular reflex system. The necessity for an adaptation occurs because of a 90 degrees displacement of the vestibular with respect to the extraocular coordinate axes during metamorphosis. Since a reorganization of vestibuloocular pathways must be hypothesized (12), the location and termination of electrophysiologically identified secondary vestibular neurons with focus on the horizontal canal system was studied with the intracellular horseradish peroxidase method in adult winter flounders. Pseudopleuronectes americanus. The oculomotor target sites of vertical canal related neurons were similar to those described in mammals. Presumed excitatory anterior canal neurons bifurcated after the main axon had crossed the midline. The descending branch headed toward the spinal cord. The ascending branch reached the oculomotor nucleus via the contralateral medial longitudinal fasciculus and terminated in the superior rectus and inferior oblique subdivisions. Presumed inhibitory posterior canal neurons ascended ipsilaterally in the medial longitudinal fasciculus and terminated mainly in the superior rectus and inferior oblique subdivisions. Horizontal canal neurons exhibited characteristics distinctly different from mammalian ones. Two types of second-order neurons were observed. In the first case, cell bodies were located in the anterior portion of the vestibular nuclear complex. After crossing the midline, the axon ascended in the contralateral medial longitudinal fasciculus. Major termination sites were found in the inferior oblique and superior rectus subdivisions of the oculomotor nucleus. Axonal branches then recrossed the midline and terminated in identical locations on the ipsilateral side. In the second case, cell bodies were located in the descending vestibular nucleus. Their axons crossed the midline and also ascended in the contralateral medial longitudinal fasciculus. Major termination sites were in the trochlear nucleus and in the inferior rectus subdivision of the oculomotor nucleus. As in the first case, axonal branches also recrossed the midline and terminated in identical motoneuron pools on the ipsilateral side. The above target sites were exactly those expected to be used in a reciprocal excitatory-inhibitory fashion during compensatory eye movements. Head-down movement would be excitatory for the lower horizontal canal producing contractions of both superior recti and inferior obliques as well as relaxation of the antagonistic inferior recti and superior obliques.(ABSTRACT TRUNCATED AT 400 WORDS)

An anatomical study of ipsilateral retinal projections in the quail using radioautographic, horseradish peroxidase, fluorescence and degeneration techniques

The topographical organization of ipsilateral retinal projections was investigated in the pigmented quail using different anatomical tracing techniques. The orthograde labeling patterns were examined after intraocular injections of (1) tritiated tracers (autoradiography), (2) horseradish peroxidase (HRP) either alone or combined with wheat germ agglutinin (WGA), (3) the fluorescent tracer rhodamine β-isothiocyanate (RITC) and (4) after unilateral retinal ablations using degeneration methods where the material was observed at both light and electron microscope levels. The different techniques provided extremely variable results with regard to demonstrating ipsilateral retinal projections. Whereas the latter could not be revealed in the radioautographic preparations, the results derived using the degeneration and HRP methods were fragmentary. The RITC method was found to be most effective and made it possible to accurately define the different ipsilateral optic contingents and their terminal arborizations within various thalamic, pretectal and tegmentomesencephalic centers. The contradictory data in the literature regarding the existence of ipsilateral retinal projections in various avian species may be explained by differences in methodology based upon the relative degrees of resolution offered by the different techniques.

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.

The projection of three extrathalamic cell groups to the cerebral cortex of the turtle Pseudemys.

Three extrathalamic subcortical inputs to the part of the cerebral cortex that is known to receive thalamic fibers in the turtle were examined in the present study. Direct projections from the locus coeruleus, the superior medial raphe nucleus, and a wide area of the basal telencephalon that lies ventromedial to the globus pallidus were demonstrated with the horseradish peroxidase method. Fluorescence histochemistry confirmed the presence of catecholamine-containing fibers in the rostral half of dorsal cortex and also demonstrated a dense network of serotoninergic fibers. Biochemical analysis showed the concentration of both monoamines to be relatively high; the norepinephrine concentration was 709 ng/g and the serotonin concentration was 1,750 ng/g. No evidence was found to suggest the existence of either a dopamine fiber projection to cortex comparable to that of mammalian neocortex or the presence of an epinephrine pathway to turtle cortex equivalent to the epinephrine-containing fibers in the pallium of amphibians. The coexistence of the projections from the thalamus with noradrenergic projections from the locus coeruleus, serotoninergic projections from the superior medial raphe nucleus, and presumably cholinergic projections from the basal telencephalon provide at least four distinct subcortical inputs to the reptilian dorsal cortex. Neither thalamic nor similar extrathalamic inputs have been demonstrated in the dorsal pallium of amphibia. Mammalian neocortex, in contrast, has even more elaborately differentiated inputs of both types. These results support the idea that thalamic and extrathalamic inputs to cortex appear at the same time in vertebrate evolution, and that both types of inputs are required for the normal development and function of neocortex.

Organization of the commissural projection to the dentate gyrus is unaltered by heavy ethanol exposure during gestation

The anterograde horseradish peroxidase method was used to determine if prenatal exposure to ethanol affected the development of the characteristic afferent lamination pattern of the commissural projection to the dentate gyrus. Mean ethanol consumption for the ethanol-consuming dams was 12.7 g/kg ± 0.3 g per day. Adult offspring of rats that consumed a liquid diet containing 35% ethanol-derived calories during days 1–21 of gestation, and both pair-fed and normal controls were examined. Brain weights and volumes of the ethanol and pair-fed control rats did not differ significantly from normal controls. However, body weights of ethanol-exposed rats were significantly reduced compared to normal controls. Computer-assisted image analysis of the HRP-labeling revealed that in spite of the heavy ethanol exposure there was no evidence of alterations in the spatial distribution of the commissural terminal field.

Organization of afferent connections to the lateral and interpositus cerebellar nuclei from the brainstem relay nuclei: a horseradish peroxidase study in the cat

Afferent projections to the lateral (dentate) and interpositus cerebellar nuclei from the brainstem relay nuclei were studied in cats using the horseradish peroxidase (HRP) method. In the first series of experiments, HRP was injected into the brachium pontis. Mossy fiber terminals were anterogradely labeled, predominantly in the lateral (hemispherical) part, moderately in the intermediate part, and slightly in the vermal part of the cerebellum. Besides these terminals in the cerebellar cortex, axon terminals labeled anterogradely were also found in the cerebellar nuclei. The labeled terminals appeared almost exclusively in the lateral nucleus and rarely in the interpositus nucleus. Cells labeled retrogradely were found both in the pontine nuclei and the tegmental reticular nucleus, but not in other brainstem nuclei. In the second series of experiments, HRP was injected into the lateral and interpositus nuclei, and retrograde labeling was examined in the brainstem relay nuclei. After HRP injection into the lateral nucleus, the number of labeled cells was significantly large in the pontine nuclei, but fairly small in the reticular or vestibular nuclei. The number of labeled cells was generally large in the inferior olive, mainly in the principal olive. After HRP injection into the interpositus nucleus, the number of labeled cells was moderate in the reticular or vestibular nuclei, but small in the pontine nuclei. The number of labeled cells in the inferior olive was also large, being distributed mainly in the accessory olives. These results indicate that the pontine nuclei and the principal olive provide major afferent inputs to the lateral nucleus, whereas the reticular nuclei, the vestibular nuclei and the accessory olives are the major afferent sources to the interpositus nucleus.

Neural pathways mediating the corneal blink reflex and Bell's phenomenon in the cat

Two central projections from the corneal representation of the sensory trigeminal complex in the cat were demonstrated with horseradish peroxidase, autoradiographic and Golgi methods: (1) to the dorsal subdivision of the ipsilateral facial nucleus that innervates the orbicularis oculi muscle; and (2) to the bordering area between the contralateral central gray matter and the oculomotor nucleus, which receives dendrites of the oculomotor cells innervating the contralateral superior rectus muscle. These two routes probably mediate early responses of the corneal blink reflex and Bell's phenomenon, respectively.

The differential projection of two cytoarchitectonic subregions of the inferior parietal lobule of macaque upon the deep layers of the superior colliculus.

Parietotectal projections were studied in the macaque monkey in experiments designed to compare the distribution of fibers originating in two cytoarchitecturally distinct regions within the inferior parietal lobule: the inferior bank of the intraparietal sulcus (area POa of Seltzer and Pandya, '80) and the adjoining part of area PG (von Bonin and Bailey, '47) on the convexity of the hemisphere, here called PGc. A dense fiber projection from POa to the intermediate and deep layers of the superior colliculus was observed by both anterograde autoradiographic and anterograde horseradish peroxidase methods. In contrast, only faint labeling was seen in the superior colliculus following injections of tritiated amino acids into area PGc on the convexity. In a second set of experiments, injections of horseradish peroxidase were placed in the intermediate and deep layers of the superior colliculus so that the cells of origin of the parietotectal projections could be identified. Many retrogradely labeled neurons were observed in POa, whereas very few labeled neurons were present in any other subdivision of the inferior parietal lobule or in the superior parietal lobule. These findings demonstrate that area POa has a prominent direct efferent projection to a major premotor region of the brainstem oculomotor system and suggest that by virtue of its parietotectal connection, this sulcal subdivision may have functional properties not shared with other subdivisions of the inferior parietal lobule.

The postnatal development of retinocollicular projections in normal hamsters and in hamsters following neonatal monocular enucleation: A horseradish peroxidase tracing study

The pattern of distribution of the retinal projections to the superior colliculus (SC) has been studied in developing normal hamsters and in hamsters following unilateral eye enucleation at birth, using the anterograde horseradish peroxidase (HRP) method. The results show that in normal hamsters the contralateral retinocollicular projection has already reached the caudal pole of the SC on the day of birth, and covered the entire SC by day 1. The ipsilateral retinocollicular projection is distributed only to the rostrolateral portion of the SC on day 0, but has covered the entire area of the SC on day 1. The innervation of the SC by the ipsilateral projection increases gradually until it reaches its maximum density on day 3 or day 4. Beginning on day 6, the density of the ipsilateral projection decreases markedly except in areas where there is a distinct clumping of retinal fibers. The normal adult pattern, which consists of dense clumps of ipsilateral retinal projections in the rostral half of the SC and a sparse ipsilateral retinal projection distributing in almost the entire extent of the SC, is established on day 10. In animals in which one eye was removed on the day of birth, the ipsilateral projection is observed in the rostral two-thirds of the SC on day 1, and innervates the entire extent of the colliculus on day 2. On day 3 or day 4, this projection is denser than that found in normal animals of the same age. The SC remains heavily innervated by ipsilateral fibers on and after day 6. The abnormal adult pattern, which consists of dense ipsilateral retinal projections in most parts of the SC, is observed on day 10. The anomalous ipsilateral retinocollicular projection which develops in eye-enucleated animals suggests that there is a competitive interaction between fibers from the two eyes during development which is critical in shaping the normal adult pattern of the ipsilateral retinocollicular projection. The early development of the contralateral projection is in advance of the ipsilateral projection, and removal of the contralateral fibers by eye enucleation at birth seems to result in a further delay in the development of the ipsilateral projection. This suggests that in the hamster the contralateral fibers may play a role in guiding the ipsilateral fibers to reach their target region. The normal development of the ipsilateral projection can be divided into two phases: there is an initial progressive increase in the density of the projection, followed by a decrease in the extent and, in some regions, the density of the projection. The implies that some of the ipsilateral fibers are transient in nature and are eliminated during development. In unilaterally enucleated hamsters the ipsilateral projection remains substantial after day 6 which suggests that some of the uncrossed fibers which would normally be eliminted are retained. In addition, the absolute extent of the enhanced ipsilateral projections is larger than the absolute extent of the normal projection at its greatest. These observations suggest that rerouting and/or sprouting of the uncrossed retinal fibers may also be important in generating the enlarged projection observed in hamsters with one eye removed at birth.

A rapid luminescently monitored enzyme immunoassay for IgE

A novel halophenol-enhanced luminol-peroxide luminescent detection method for horseradish peroxidase has been tested in an enzyme immunoassay for IgE. The luminescent enyzme immunoassay was reproducible (within-batch CV, 3.9–13.2%) and values obtained on serum samples showed good agreement with those obtained by colorimetric enzyme immunoassay. The major advantages of the luminescent detection method are that it is rapid, 30 s compared with 30 min for the colorimetric assay using o-phenylenediamine, and the luminescent signal is intense and stable for several minutes.

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.

An EM-autoradiographic and EM-HRP study of the commissural projection of the superior colliculus in the cat.

Terminals of the commissural projection in the cat were characterized ultrastructurally by autoradiographic and horseradish peroxidase methods. The results of the two studies are complementary. Terminals of commissural cells are present in the intermediate and deep layers of the cat superior colliculus. Two distinct populations of terminals are present: one containing mostly round vesicles and forming asymmetric specializations, and a second containing mostly pleomorphic vesicles and forming symmetric specializations. Both populations contact small dendrites or dendritic appendages. The two populations, mostly round and mostly pleomorphic, are present in the ratio of 2:1. Terminals measure approximately 1.1 micron in mean diameter and contact profiles ranging in size from 0.2 to 4.6 micron. There is no significant difference between the two populations in either pre- or postsynaptic profile size. The colocalization of terminals of commissural neurons with other afferent and efferent projections of the intermediate layers of the superior colliculus is discussed.

Direct cortical projections to the parabrachial nucleus in the cat.

Direct projections from the cerebral cortex to the parabrachial nucleus in the cat were examined by the horseradish peroxidase (HRP)method. When HRP was injected into the parabrachial nucleus, retrogradely labeled neuronal cell bodies were seen, bilaterally with an ipsilateral predominance, mainly in the orbital gyrus, the lateral bank of the presylvian sulcus, and a restricted region in the infralimbic cortex on the medial surface of the frontal lobe (stereotaxic coordinates; Fr: 22, L: 1, H: -1); all labeled neurons were in deep pyramidal cell layer. After injecting HRP conjugated to wheat germ agglutinin (WGA-HRP) into the cortical regions where retrogradely labeled neurons were found after injecting HRP into the parabrachial nucleus, anterogradely labeled cortical fibers were traced to the parabrachial nucleus. Corticoparabrachial fibers originating from the orbital gyrus and the lateral bank of the presylvian sulcus ran ipsilaterally through the internal capsule and the cerebral peduncle down to the lower brainstem, whereas those from the infralimbic cortex coursed down ipsilaterally through the medial forebrain bundle. These cortical fibers to the parabrachial nucleus were distributed bilaterally with an ipsilateral predominance. Cortical fiber terminals in the parabrachial nucleus were topographically arranged: Corticoparabrachial fibers from the lateral bank of the presylvian sulcus ended most massively in the dorsal part of the lateral parabrachial nucleus. Corticoparabrachial fibers from the orbital gyrus ended most heavily in the medial parabrachial nucleus and less heavily in the lateral parabrachial nucleus. Corticoparabrachial fibers from the infralimbic cortex ended mostly in the parabrachial regions surrounding the brachium conjunctivum.

Cerebellar, medullary and spinal afferent connections of the paramedian reticular nucleus in the cat

The topographic organization of afferent projections from the deep cerebellar nuclei, medulla oblongata and spinal cord to the paramedian reticular nucleus (PRN) of the cat was studied using the horseradish peroxidase (HRP) method of retrograde labelling. Discrete placements of HRP within each of the dorsal (dPRN) and ventral (vPRN) regions of the PRN showed some segregation of input. The deep cerebellar nuclei project in a predominantly contralateral fashion upon the PRN. A small but significant ipsilateral fastigial afferent component is also present. The fastigial and dentate nuclei contribute the majority of fibers to the dPRN whereas the interposed nucleus provides very little. The vPRN receives a relatively uniform input from all 3 cerebellar nuclei. Both lateral vestibular nuclei contribute the majority of fibers from the vestibular nuclear complex largely from their dorsal division. Additional input arises from bilateral medial and inferior vestibular nuclei. The vPRN receives relatively more fibers from the inferior vestibular nuclei than does the dPRN while inputs from the medial vestibular nuclei are comparably sparse. The PRN receives bilateral projections from the nucleus intercalatus (of Staderini). A significant projection to the contralateral PRN occurs from the ventrolateral subnucleus of the solitary complex and its immediate vicinity. Additional sources of medullary afferent input include the lateral, gigantocellular and magnocellular tegmental fields, the contralateral PRN and the raphe nuclei. Sites of origin of spinal afferents to the dPRN are bilaterally distributed mainly within Rexed's laminae VII and VIII of the cervical cord whereas those to the vPRN are confined largely to the medial portion of the contralateral lamina VI in the C1 segment. A few labelled cells are found in the thoracolumbar cord with those to the vPRN being more caudal. These data provide the neuroanatomical substrate for a better understanding of the functional role of the PRN in mediating cardiovascular responses appropriate to postural changes.

Detection of caldesmon in muscle and non-muscle tissues of the chicken using polyclonal antibodies

Polyclonal antibodies raised in rabbits against chicken gizzard caldesmon have been purified and used in immunoblotting experiments to study the distribution of this actin- and calmodulin-binding protein in diverse tissues of the chicken. Total homogenates and heat-treated homogenate supernatants derived from each tissue were subjected to sodium dodecyl sulfate-polyacrylamide gradient slab gel electrophoresis and immunoblotting using the horseradish peroxidase method. All chicken tissues examined contained caldesmon of Mr = 141,000. The amount of caldesmon found in the different tissues varied considerably and semi-quantitative comparison of stained immunoblots indicated the following relative caldesmon contents: gizzard greater than oesophagus greater than duodenum = small intestine greater than lung greater than aorta greater than heart = skeletal muscle greater than kidney = trachea greater than brain greater than liver. Each tissue revealed small amounts of lower Mr immunoreactive proteins, predominantly bands of Mr 94,000 and 70,000, which appear to be proteolytic fragments of caldesmon. Isolated caldesmon was found to be highly sensitive to proteolysis. The widespread distribution and similarity of caldesmon in different tissues of the chicken suggest its functional importance and structural conservation.