Ascending and descending cholinergic fibers in cat spinal cord: histochemical evidence.

The distribution of fibers staining for cholinesterase in the spinal cord of the cat was examined after hemisection at the level of the third cervical segment (C3), of the tenth thoracic segment (Ti 0), or of the first lumbar segment (LI). An accumulation of cholinesterase was found in many fibers of the cord both rostral and caudal to the lesion, the distribution being different in the two regions. These experiments indicate that there are ascending and descending cholinergic fibers in cat spinal cord.

A light and electron microscopic study of the inferior olivary nucleus of the squirrel monkey, Saimiri sciureus.

This study provides a description of the normal morphology of the inferior olive of the squirrel monkey, Saimiri sciureus, at the light and electron imcroscopic level. The cytoarchitecture of the inferior olive was maped from serial transverse sections stained with cresyl violet. In common with other mammals, the inferior olive of the squirrel monkey consists of three subdivisions. The medial accessory olive includes seven subnuclei. Both the dorsal and medial accessory olives extend through approximately 90% of the total length of the inferior olivary complex. The principal olive, consisting of a dorsal and ventral lamella continuous with one another laterally, extends through the rostral 55% of the inferior olive. It is somewhat less convoluted than the principal olive of the macaque (Bowman and Sladek, '73). In most other respects, the inferior olive of the two primates is quite similar. Two patterns of dendritic arborization are noted in Golgi preparations from the caudal principal and accessory olives. Dendrites streaming away from the soma, and dendrites curling around the soma in a "ball-like" pattern were observed in all three subdivisions of the inferior olive caudally. Simple spines are occasionally seen on the soma, and a few simple or club-shaped spines were noted on the proximal portion of the dendritic arborization. Spines are more numerous on distal portions of the dendritic tree, however, and include simple, filiform, club-shaped and occasionally complex, or racemous, spiny appendages. Viewed in the electron microscope, most inferior olivary neurons are seen to contain the typical organelles with the usual conformation and distribution. Rarely, a neuron with an indented nucleus and a thin rim of cytoplasm containing a paucity of organelles and a wispy endoplasmic reticulum is encountered. Axon terminals containing either clear round or clear pleomorphic vesicles are seen in all three olivary subdivisions. In a random survey of 706 axon terminals, 54% contained predominantly clear round vesicles. Large dense cored vesicles are seen in varying numbers in both types of terminals. Rarely, profiles containing mainly large dense cored vesicles are ob served. Axosomatic synapses involving both types of clear vesicle containing terminals are occasionally encountered. Such synapses are symmetrical, regardless of the type of vesicle found in the axon terminal. Axodendritic synapses involving round vesicle containing terminals are assymetrical, while those involving pleomorphic vesicle containing terminals are usually, but not invariably, symmetrical. Axondendritic synapses occur at all levels of the dendritic tree. Very rarely, synapses between an axon terminal and a profile resembling a dendrite, but containing pleomorphic vesicles, has been observed. Synaptic clusters, consisting of a central core of small dendritic elements surrounded by both round and pleomorphic vesicle containing terminals, are found in all three subdivisions of the inferior olive...

A light and electron microscopic study of the interstitial nucleus of Cajal in rat.

The morphology of the interstitial nucleus of Cajal (INC) in the rat was studied with the light and electron microscope. The INC was mapped throughout its rostrocaudal extent from cresyl violet-stained frozen sections cut transversely through the midbrain in the stereotaxic plane. Caudally, the INC consisted of a small number of scattered cells lying ventrolateral to the periaqueductal grey. In three of four cases studied, the caudal tip of the nucleus was located between 40 and 120 micrometers rostral to the rostral tip of the somatic cell columns of the oculomotor nucleus. Proceeding rostrally, the INC increased in size, reaching its maximal development just caudal to its most rostral extent. The INC was limited rostrally by the fibers of the fasciculus retroflexus. The mean rostrocaudal length of the INC was 1.12 mm. On the basis of light microscopic observations of cresyl violet-stained paraffin sections, two groups of neurons could be distinguished in the INC. One group consisted of large, oval to multipolar cells with mean dimensions of 33 X 23 micrometers. The second group, which included by far the greatest number of cells, consisted of small to medium neurons, round, triangular, polygonal or fusiform in shape, with mean dimensions of 19 X 14 micrometers. Injection of horseradish peroxidase into lesions in the cervical spinal cord resulted in retrograde labeling of neurons of all sizes and shapes throughout the length of the INC. Labeled neurons were also found in the red nucleus, the mesencephalic reticular formation, and the nucleus of the posterior commissure. All the morphological varieties of neurons described with the light microscope could be identified in the electron microscope. Large neurons, and some cells of the small to medium group, contained well developed Nissl bodies together with numerous cytoplasmic organelles. Many neurons in the small to medium group, however, did not contain conspicuous Nissl bodies, and had a poorly developed rough endoplasmic reticulum. Axon terminals containing either round or pleomorphic vesicles were seen in the INC. Axosomatic synapses were formed by both types of terminals. Such synapses were usually symmetrical, regardless of the shape of the vesicles within the terminal. In a number of neurons, the percentage of the surface of the neuronal somata in direct apposition to axon terminals was measured. The results of such measurements suggest that a greater percentage (more than 50%) of the surface of larger neurons is apposed by axon terminals than is the case with smaller neurons, which, on the average, were invested by axon terminals over 15% of their total surface in any given single plane of section. Axon terminals investing the surfaces of proximal dendrites were morphologically similar to those in apposition to neuronal somata.

Cytoarchitecture, neuronal morphology, and some efferent connections of the interstitial nucleus of Cajal (INC) in the cat.

This study describes the cytoarchitecture and neuronal morphology of the interstitial nucleus of Cajal (INC) in the cat. In addition, the efferent projections of this nucleus to the spinal cord and inferior olive were studied by retrograde labelling with horseradish peroxidase (HRP). The INC was shown to extend rostrocaudally for slightly more than 2 mm. Caudally, the nucleus consists of a small number of loosely aggregated neurons lying lateral to the ventral periaqueductal gray matter at a rostrocaudal level corresponding to the rostral one-fifth of the somatic cell columns of the oculomotor nucleus. Rostrally, the INC increases in size and reaches its maximum development in its rostral half, where it lies ventrolateral to the nucleus of Darkschewitsch (ND). Rostrally the INC is bounded by the dorsoventrally aligned fibres of the fasciculus retroflexus. Two groups of neurons could be distinguished within the INC in both normal and HRP-injected material. One group consists of a relatively small number of large, oval, pyramidal, fusiform, or multipolar neurons with mean dimensions of 40 X 26 micrometers. The second group consists of numerous small to medium-sized neurons with mean dimensions of 20 X 14 micrometers. Large neurons and some cells of the second group contain substantial amounts of Nissl substance throughout their perikarya. Some medium-sized to small neurons exhibit indentations in their nuclei, and glial cells are often apposed to their cell membranes. Golgi-Kopsch preparations taken from kitten showed that INC neurons possess sparsely branched, radiating dendritic trees with few spinous processes. The majority of INC neurons retrogradely labelled with HRP exhibited similar dendritic patterns. Injections of HRP into lesions at cervical, thoracic, or lumbar levels of the spinal cord resulted in retrograde labelling of neurons of all sizes and shapes throughout the entire length of the INC. However, the greatest number of HRP-labelled cells in INC were observed subsequent to injections of the enzyme into cervical levels of the cord. Following injections of HRP into the inferior olive only small to medium-sized neurons were labelled in the nucleus, the majority of which are located in rostral levels of the INC. A substantial olivary projection was observed to originate in the nucleus of Darkschewitsch (ND) and the nucleus parafascicularis (NPF). The sizes of the projections from these two nuclei to the inferior olive appeared to be much larger than that from the INC. Smaller numbers of neurons were also observed in the rostral parvocellular red nucleus (RN) and mesencephalic reticular formation (MRF).

Observations on the afferent and efferent organization of the vagus nerve and the innervation of the stomach in the squirrel monkey.

Horseradish peroxidase was injected into the cervical vagus nerve or stomach wall of adult squirrel monkeys. Following cervical vagus nerve injections, labelled afferent fibres were present in the tractus solitarius and labelled fibres and terminals were present in medial and lateral parts of the nucleus of the tractus solitarius (NTS) ipsilaterally. Afferent labelling was also seen in the ipsilateral commissural nucleus and in the area postrema. Labelling was present contralaterally in caudal levels of the medial parts of the NTS, in the commissural nucleus, and in the area postrema. Afferent projections to the ipsilateral pars interpolaris of the spinal trigeminal nucleus and to the substantia gelatinosa of the C1 segment of the spinal cord were also labelled. Following injections of HRP into the anterior and posterior stomach walls, the tractus solitarius was labelled bilaterally. Afferent labelling was concentrated bilaterally in the dorsal parts of the medial division of the NTS, i.e., in the subnucleus gelatinosus, and in the commissural nucleus. The regions of NTS immediately adjacent to the tractus solitarius were largely unlabelled. Injections of HRP into the cervical vagus nerve resulted in heavy retrograde labelling of neurons in the ipsilateral dorsal nucleus of the vagus (DMX) and in the nucleus ambiguus (NA). In addition a few neurones were labelled in the intermediate zone between these two nuclei. Retrogradely labelled neurons were also present in the nucleus dorsomedialis in the rostral cervical spinal cord and in the spinal nucleus of the accessory nerve. Injections of HRP into the left cricothyroid muscle in two cases resulted in heavy retrograde labelling of large neurons in the left NA. Following stomach wall injections of HRP retrograde labelling of neurons was seen throughout the rostrocaudal and mediolateral extent of the DMX; there was no apparent topographical organization of the projection. In these cases, a group of labelled smaller neurons was found lying ventrolateral to the main part of the NA through its rostral levels. This study in a primate indicates that a large vagal afferent projection originates in the stomach wall and terminates primarily in the subnucleus gelatinosus of the NTS and in the commissural nucleus with a distribution similar to that described previously in studies in several subprimate mammalian species. The present results and those of other studies suggest some degree of segregation of visceral input within different subnuclei of the NTS.(ABSTRACT TRUNCATED AT 400 WORDS)

An ultrastructural study of the red nucleus in the rat.

The red nuclei of 14 adult male rats of the Wistar strain were prepared for electron microscopic study following perfusion with a mixture of aldehydes, Neurons of four size categories were identified in 1 mu Epon sections and their ultrastructural characteristics were studied in adjacent thin sections. Giant (greater than 40 mu) and large (26-40 mu) neurons are distinguished primarily by size and possess similar ultrastructural features: extensive areas of rough endoplasmic reticulum (RER), a prominent perinuclear Golgi complex, numerous mitochondria and pigment granules and a large, ovoid nucleus which occasionally contains intranuclear rodlets. Medium size neurons (20-25 mu) have less extensive, poorly organized RER and randomly distributed Golgi complexes. The nuclear envelopes of these cells frequently show multiple invaginations and continuity with the RER cisternae. In small neurons (less than 20 mu) the RER occurs as single or anastomosing strands whi le golgi complexes and pigment granules are few. In both medium size and small neurons, aggregates of condensed chromatin are adherent to the inner nuclear membrane. Three main types of synaptic terminals may be distinguished in the red nucleus: (1) small terminals with flattened vesicles and symmetrical densities (F terminals), (2) small terminals with rounded vesicles and asymmetrical densities (RS terminals), and (3) large (10-15 mu) asymmetrical, rounded vesicle terminals which form multiple contacts along their length (RL terminals). The small neurons receive both F and RS terminals on their dendrites and infrequently on their cell somas. The large and giant neurons receive F, RS and RL terminals on their somas and proximal dendrites and F and RS terminals on their distal dendrites. The somas and dendrites of medium size neurons receive both F and RS terminals but RL terminals do not lie in relation to them. Spine contacts are common throughout the nucleus and occur on both somas and dendrites.

A reevaluation of midbrain and diencephalic projections to the inferior olive in rat with particular reference to the rubro-olivary pathway.

Projections from the midbrain and caudal diencephalon to the inferior olivary nucleus (ION) in the rat were investigated by using anterograde and retrograde tracing techniques. Particular attention was directed to studying the projection from the red nucleus (RN) to the ION. Tritiated leucine was stereotaxically injected into the RN in six animals. The injection sites and the portion of the medulla containing the ION were processed for autoradiography. In three cases, the injections were largely confined to the RN. In these instances, no terminal labeling was noted in the ION. In the other three animals, the injections spread beyond the RN to include, in one or another of the rats, such areas as the mesencephalic reticular formation (MRF) between the RN and the periaqueductal gray (PAG), the region around the fasciculus retroflexus (FR), the prerubral area, the MRF dorsolateral to the RN, or the region dorsal to the medial geniculate body. In these animals, terminal labeling was observed ipsilaterally in the ION, chiefly in the caudal one-half of the medial accessory olive and in the dorsal lamella and lateral bend of the principal olive. Retrograde transport of lectin-conjugated horseradish peroxidase (WGA-HRP) or the fluorescent dye fast blue (FB), injected into the ION, was used to localize regions in the midbrain and caudal diencephalon containing neurons which project to the inferior olive. Following WGA-HRP injections into the ION, retrogradely labeled neurons were found in the ipsilateral ventral PAG, the MRF between the PAG and the RN, the MRF dorsolateral to the RN, and the area dorsal to the medial geniculate body. Large numbers of heavily labeled neurons were found in the area surrounding the FR. No labeled cells were noted in the RN or the nucleus of Darkschewitsch (ND). Subsequent to FB injections into the ION, fluorescent neurons were observed in the same regions which contained HRP-labeled neurons in the experiments using the enzyme as a tracer. Additionally, very small numbers of FB-labeled neurons were found in the RN and the ND. These results indicate that, in the rat, descending input from the midbrain and caudal diencephalon to the ION arises chiefly from a medially located column of neurons extending from the level of the rostral RN to the region of the FR. Little, if any, input to the ION is derived from the ND or RN in this species, unlike the cat, in which a substantial projection to the ION arises from the ND and the RN.

The ultrastructure of the subnucleus gelatinosus of the nucleus of the tractus solitarius in the cat.

The dorsomedial region of the nucleus of the tractus solitarius termed the subnucleus gelatinosus (SNG) was studied at the light and electron microscopic level in the cat. In cresyl violet and luxol fast blue stained sections the SNG contained small neuronal somata that were scattered throughout a pale-staining neuropil containing few myelinated fibers. These neurons were difficult to impregnate with Golgi staining techniques, but in successful impregnations the somata were observed to be 10--19 micrometers in diameter and bore few sparsely branching primary dendrites. Spines were present on the dendrites of some neurons and were more numerous on distal portions of the dendritic tree. Ultrastructural examination of the SNG revealed that the neuronal complement consisted of round, oval, or spindle shaped neurons with little or no organized Nissl substance. Rare myelin-like ensheathments of neuronal perikarya were also observed. Bundles of fine unmyelinated axons that coursed mainly longitudinally were a prominent feature of the area. The most common type of axon terminal observed contained mainly round clear vesicles, approximately 31 nm in diameter, and made asymmetrical synaptic contact with a dendritic profile. Pleomorphic vesicle-containing terminals involved in symmetrical synaptic contact were also commonly seen. Axodendritic and axosomatic synapses were associated with terminals containing either round clear vesicles or pleomorphic vesicles. Less commonly, dendrodendritic and dendrosomatic synapses were seen, the presynaptic elements of which contained pleomorphic vesicles. Following removal of a nodose ganglion, degenerating terminals of vagal afferent fibers were observed throughout the neuropil. Such terminals contained round, clear vesicles with an occasional large, dense-cored vesicle, and made axodendritic and axosomatic synaptic contacts.

The central distribution of vagal catecholaminergic neurons which project into the abdomen in the rat.

A double labeling technique employing retrograde labeling of vagal neurons with horseradish peroxidase from injections into the stomach wall and immunocytochemistry for dopamine-beta-hydroxylase revealed catecholaminergic neurons in the medulla oblongata which project into the abdomen. The great majority of such neurons were located in the dorsal motor nucleus of the vagus, particularly in its rostral third.

Acetylcholinesterase activity and acetylcholine effects in the cerebello-rubro-thalamic pathway of the cat.

Unilateral transections of the brachium conjunctivum (BC) of cats resulted, after 2-3 weeks, in marked loss of acetylcholinesterase (AChE) activity from the contralateral red nucleus (RN) and ventral tier nuclei of the thalamus (VA-VL). Significant changes in activity were not observed in other locations. Sensitivity of RN neurons to iontophoretically applied acetylcholine (ACh) was studied under conditions which should maximize ACh sensitivity, including AChE inhibition, but ACh was found to have only a weak depressant effect on excitability or no effect at all. Intravenous physostigmine usually increased spontaneous activity of RN neurons, and sometimes increased potentials evoked by electrical stimulation of cerebellar nuclei, to a small extent. Anticholinergic drugs were found not to influence such evoked responses, except to reverse the effects of physostigmine. It is concluded that ACh is not a major transmitter in the excitatory cerebello-rubral tract in spite of the relationship of AChE to this pathway.

Projections from the interstitial nucleus of Cajal to the inferior olive and to the spinal cord in cat: a retrograde fluorescent double-labeling study.

Injections of the fluorescent dyes Fast Blue (FB) and Nuclear Yellow (NY) were placed in the inferior olive and cervical spinal cord respectively in three experimental animals. Results showed that the interstitial nucleus of Cajal (INC) projected mainly to the spinal cord, with only a modest termination within the inferior olive. The nucleus of Darkschewitsch and the rostromedial portion of the red nucleus projected heavily to the inferior olive but not to the spinal cord. Very few INC neurons were double-labeled with FB and NY, suggesting that only a small minority of spinal projecting neurons in the INC give rise to collaterals which terminate within the inferior olive.

Gastric afferents to the nucleus of the solitary tract in the cat.

Afferent projections of the vagus nerve (X) were investigated following nodose ganglion injections of [3H]leucine or horseradish peroxidase (HRP) and stomach wall injections of HRP. Ganglionic injections demonstrated a projection to the solitary nucleus (SN) including the part designated as the area subpostrema (ASP). A projection to the area postrema was also seen following [3H]leucine injections. Stomach wall injections of HRP labelled fibres and terminals bilaterally in the ASP and more ventral parts of the SN. These results provide the first morphological evidence of gastric afferents of X projecting to an area of the SN previously implicated in gastric functions.

An electron microscopic study of the direct spino-olivary projection to the inferior olivary nucleus in the rat.

Prior to the electron microscopic identification of spino-olivary terminals, the distribution of the direct spino-olivary projection was determined at the light microscopic level. Lesions were placed in the cervical or thoracic spinal cord in 7 rats. After a 7 day survival, animals were perfused, and transverse sections through the medulla were processed according to the Fink-Heimer technique. Terminal degeneration was found in the caudal third of the medial accessory olive, and in the lateral half of the dorsal accessory olive throughout its length. In order to determine the ultrastructural characteristics of direct spino-olivary terminals, lesions were placed in the cervical spinal cord of a further 4 rats. After a 2 or 3 day survival period, the animals were processed for electron microscopy. The portions of the inferior olivary nucleus in which degeneration had been noted in the Fink-Heimer studies were sampled and examined in the electron microscope. The majority of degenerating spino-olivary terminals underwent an electrondense change. Most degenerating terminals contained round, clear vesicles, but a few were found to contain predominantly flattened vesicles. Degenerating terminals contacted small dendritic profiles or spines, neither of which were associated with synaptic clusters.

The ultrastructural identification of vagal terminals in the solitary nucleus of the cat after anterograde labelling with horseradish peroxidase.

Horseradish peroxidase (HRP) was injected into the nodose ganglion of the cat in order to label the axon terminals of vagal afferent fibres in the solitary nucleus. HRP reaction product was seen in axons, the majority of which were unmyelinated. Axon terminals containing round clear vesicles and an occasional dense-core vesicle were also labelled. These terminals synapsed upon both large and small dendritic elements, the majority of which were less than 1.0 micrometers in diameter. The labeling method appears to affect adversely the normal ultrastructural characteristics of terminals or their postsynaptic relationships and in these respects is superior to electron microscopic degeneration methods.

Projections between the hypothalamus and the dorsal vagal complex in the cat: an HRP and autoradiographic study.

The hypothalamus is known to be intimately involved in the control of autonomic function. This study provides detailed information about pathways between the hypothalamus and the dorsal vagal complex in cat. Injection of horseradish peroxidase into the dorsomedial medulla produced retrograde neuronal labeling in the paraventricular nucleus of the hypothalamus. Injection of 3H-leucine into the paraventricular nucleus produced dense anterograde labeling in the dorsal motor nucleus of the vagus, and lighter labeling in the nucleus of the tractus solitarius, particularly in its medial subnucleus. The subnucleus gelatinous was virtually free of label, except in its medial and lateral portions. Anterograde labeling was distributed bilaterally, with an ipsilateral predominance. Injection of horseradish peroxidase into the area of the paraventricular nucleus produced retrograde neuronal labeling bilaterally in the nucleus of the tractus solitarius and the reticular formation ventrolateral to the dorsal vagal complex. anterograde terminal labeling overlapped the distribution of retrogradely labeled neurons. These findings are compared to those in rat, and discussed in relation to their functional implications.

Cortical projections to the nucleus of the tractus solitarius: an HRP study in the cat.

Recent evidence suggests that autonomic reflexes involving sensations such as olfaction and gustation may be cortically mediated via centripetal pathways to brainstem autonomic centers. A study was therefore undertaken to elucidate one of these pathways in greater detail. Lectin conjugated horseradish peroxidase was injected into the nucleus tractus solitarius. Following standard light microscopic histochemical procedures to reveal horseradish peroxidase activity, the distribution of retrogradely labeled neurons in the cortex was recorded. Retrogradely labeled somata were seen bilaterally in layer five of the orbital gyrus, anterior insular cortex and infralimbic cortex. In other cats, the same tracer was injected into the orbital gyrus or anterior insular cortex. Bilateral anterograde labeling was seen in various subnuclei throughout the rostrocaudal extent of the nucleus tractus solitarius, but was heaviest in rostral regions of the nucleus. Labeling was also seen bilaterally in the spinal trigeminal nucleus. The projection to the nucleus tractus solitarius could allow for cortical modulation of gustatory and visceral information which is conveyed to the brainstem via the facial, glossopharyngeal and vagus nerves.

Direct vagal input to neurons in the area postrema which project to the parabrachial nucleus: an electron microscopic-HRP study in the cat.

This study in cat examines the synaptic relationship of vagal afferents to parabrachial projecting neurons in the area postrema (AP) using anterograde and retrograde transport of horseradish peroxidase (HRP). Wheat germ agglutinin-HRP injected into the parabrachial nucleus (PBN) produced retrograde neuronal labeling in the AP and in the nucleus of the tractus solitarius bilaterally, but with an ipsilateral predominance. Labeled neurons were confined mainly to the caudal 2/3's of the AP. Following injection of WGA-HRP into the PBN and HRP into the nodose ganglion in the same animal, examination of sections of the AP with the electron microscope revealed anterogradely labeled axon terminals in apposition to retrogradely labeled somata and dendrites. In some instances, labeled terminals were observed to form synaptic contacts with retrogradely labeled neurons. We conclude that in the cat a vagal input to neurons in the AP is monosynaptically relayed to the PBN.

A comparison of the distribution of the cerebellar and cortical connections of the nucleus of Darkschewitsch (ND) in the cat: a study using anterograde and retrograde HRP tracing techniques.

Bidirectional transport of lectin conjugated horseradish peroxidase was employed to investigate the relative distribution of the cerebellar and cortical connections of the nucleus of Darkschewitsch in the cat. Injection of horseradish peroxidase into the deep cerebellar nuclei produced terminal labeling which extended throughout the length of the contralateral nucleus of Darkschewitsch and into the perifascicular region. Injection of horseradish peroxidase into the pericruciate cortex produced both ipsilateral terminal labeling which extended throughout the length of the nucleus of Darkschewitsch and into the perifascicular region, and ipsilateral retrograde neuronal labeling. Labeled neurons displayed a variety of shapes and sizes, were more numerous in sections cut at rostral levels of the nucleus of Darkschewitsch, and were located both within and outside fields of terminal labeling. Comparison of the distribution of labeling following cerebellar and cortical injections indicates that convergence and overlap of input from these two sources occur in the nucleus of Darkschewitsch. These findings provide the morphological basis for integration of cerebellar and cortical information in this nucleus which may, in turn, influence output from neurons which project to the cortex or to the inferior olivary nucleus.