Irena Grofová

Origin and distribution of glutamate decar☐ylase in substantia nigra of the cat

The topographical distribution of glutamate decar☐ylase (GAD) in substantia nigra in unoperated and operated cats was studied in samples microdissected from freeze-dried tissue sections. The concentration of GAD, the enzyme synthesizing γ-aminobutyric acid (GABA), was highest in the medial part of pars reticulata, and decreased in the mediolateral direction. In pars compacta, on the other hand, the highest enzyme activity was found in the lateral part which merges with pars reticulata, and it decreased gradually in the latero-medial direction. The activity of GAD was always lower in the medial part of pars compacta, which contains the highest concentration of cell bodies. GAD in substantia nigra decreased after lesions in putamen, nucleus caudatus, globus pallidus and nucleus entopeduncularis. The loss of enzyme activity was strictly localized and was related to the site of termination of the degenerating striato-nigral fibers. The reduction of GAD in substantia nigra following lesions of globus pallidus or nucleus entopeduncularis may be ascribed to the interruption of striato-nigral fibers passing through these regions. The results thus indicate that the fibers of the GAD-containing axon terminals in substantia nigra of the cat originate in putamen and nucleus caudatus. Subcellular fractionation showed that about 85% of GAD and about 25% of lactate dehydrogenase were present in particles (probably synaptosomes) from substantia nigra in unoperated animals. Electron microscopic examination revealed that 11.5% of the tissue volume of pars reticulata was occupied by boutons compared to 5.9% for pars compacta. The concentration of GABA in pars reticulata was found to be 9 mM. From these data the intraterminal concentration of GABA was estimated to be at least 60 mM, probably over 100 mM. DOPA decar☐ylase was mainly found in pars compacta. Acetylcholin-esterase showed a very high activity in substantia nigra, the highest concentration being found in the medial part of pars reticulata. In contrast, the concentration of choline acetyltransferase was very low. The ratio of acetylcholinesterase activity to choline acetyltransferase activity was 1000. DOPA decar☐ylase and the cholinergic enzymes were little affected by the above described lesions.

Distribution of glutamate decarboxylase, choline acetyl-transferase and aromatic amino acid decarboxylase in the basal ganglia of normal and operated rats. Evidence for striatopallidal, striatoentopeduncular and striatonigral GABAergic fibres.

Abstract The topographical distribution of glutamate decarboxylase (GAD), aromatic amino acid decarboxylase (AAD) and choline acetyltransferase (ChAT) were studied in striatum (i.e. caudate-putamen), globus pallidus, nucleus entopeduncularis and substantia nigra. There were only small differences in the rostrocaudal distribution of enzymes in striatum. The highest concentration of GAD was found in the ventrocaudal part, whereas AAD was highest in the rostral part. ChAT did not show any distinct distribution pattern. Globus pallidus and nucleus entopeduncularis were similar in their content of GAD, AAD and ChAT. They were rich in GAD but poor in AAD and ChAT. The highest concentration of GAD was found in the rostralmost part of globus pallidus. In substantia nigra AAD was concentrated in the rostral part; contents of GAD and ChAT did not differ distinctly in the rostral and caudal nigra. GAD and ChAT were highly localized in a particulate fraction, probably in all 4 regions, whereas AAD was localized in particulate fraction in the striatum and globus pallidus and soluble in the nucleus entopeduncularis and substantia nigra. Transverse and/or oblique hemitransections which passed through the striatum but not through the globus pallidus resulted in substantial loss of GAD in the globus pallidus, nucleus entopeduncularis and substantia nigra. There was a good correlation between the anteroposterior level of hemitransection and the decline in GAD activities, which was generally highest following posterior lesions. The reduction of GAD was largest in the globus pallidus and smallest in the substantia nigra, in which a significant loss of GAD occurred only following transections of the postcommissural part of caudate-putamen. A very high decrease of GAD in the nucleus entopeduncularis and substantia nigra was obtained following posterior oblique hemitransections which passed through posterior part of striatum and rostral globus pallidus. The results indicate that the majority of GABAergic terminals in the globus pallidus belong to striatopallidal fibers. They suggest, furthermore, that a large number of striatoentopeduncular and striatonigral fibers are GABAergic, the latter arising preferentially from the posterior part of caudate-putamen.

Mesencephalic and diencephalic afferents to the superior colliculus and periaqueductal gray substance demonstrated by retrograde axonal transport of horseradish peroxidase in the cat.

The mesencephalic and diencephalic afferent connections to the superior colliculus and the central gray substance in the cat were examined by means of the retrograde transport of horseradish peroxidase (HRP). After deep collicular injections numerous labeled cells were consistently found in the parabigeminal nucleus, the mesencephalic reticular formation, substantia nigra pars reticulata, the nucleus of posterior commissure, the pretectal area, zona incerta, and the ventral nucleus of the lateral geniculate body. A smaller number of cells was found in the inferior colluculus, the nucleus of the lateral lemniscus, the central gray substance, nucleus reticularis thalami, the anterior hypothalamic area, and, in some cases, in the contralateral superior colliculus, Forels field, and the ventromedial hypothalamic nucleus. Only the parabigeminal nucleus and the pretectal area showed labeled cells following injections in the superficial layers of the superior colliculus. In the cats submitted to injections in the central gray substance, labeled cells were consistently found in the contralateral superior colliculus, the mesencephalic reticular formation, substantia nigra parts reticulata, zona incerta and various hypothalamic areas, especially the ventromedial nucleus. In some cases, HRP-positive cells were seen in the nucleus of posterior commissure, the pretectal area, Forels field, and nucleus reticularis thalami. A large injection in the mediodorsal part of the caudal mesencephalic reticular formation, which included the superior colliculus and the central gray substance, resulted in numerous labeled cells in nucleus reticularis thalami. The findings are discussed with respect to the suggested functional division of the superior colliculus into deep and superficial layers. Furthermore, the possible implications of labeled cells in zona incerta and the reticular thalamic nucleus are briefly discussed.

An experimental electron microscopic study on the striatonigral projection in the cat.

SummaryThe ultrastructure of the cats substantia nigra was investigated from 2–21 days following large lesions of the caudate nucleus and the putamen. From 4 days on a large number of degenerating boutons and degenerating unmyelinated fibers are seen in the substantia nigra, in the pars compacta as well as the pars reticulata. Both parts, mainly the latter, receive striatal afferents. The degeneration in the substantia nigra following striatal lesions is of the dark type. Most of the degenerating boutons apparently are of the type I (see Rinvik and Grofová, 1970) and end on all parts of the nigral cell surface, including the dendritic spines. One instance of a degenerating presynaptic bouton in an axo-axonic synapse has been found. Some degenerating boutons also probably belong to the type II bouton, while degenerating boutons of type III were never seen following the striatal lesions. The electron microscopic identification of early axonal degeneration in the central nervous system, is discussed with reference to the paper of Cohen and Pappas (1969). Problems concerning the pars compacta versus the pars reticulata of the substantia nigra are taken up. The possible sources of origin of the different types of boutons in the cats substantia nigra, is discussed.

Observations on the fine structure of the substantia nigra in the cat

SummaryA light and electron microscopical investigation has been undertaken of the substantia nigra in the normal cat. The pars reticulata partly contains the arborization of dendrites whose cell bodies are located in the so-called pars compacta. There is a considerable overlap of the dendritic fields in the rostrocaudal direction, while the dendritic fields are very restricted in the mediolateral extension of the substantia nigra. The secondary and all subsequent branches of the dendrites of nigral cells are for considerable distances completely covered by boutons. Only few boutons contact the cell bodies. Three types of boutons are distinguished in the substantia nigra in the cat. Type I, about 90 % of the total, is of the terminal type, contains pleomorphic vesicles and establishes symmetrical synapses with nigral cell soma, dendritic trunks and spines. The type II bouton (about 10 % of the total number) is most commonly of the terminal type, contains spherical vesicles and establishes asymmetrical synapses with cell bodies and dendritic trunks of nigral cells. The type III bouton (about 2 % of the boutons) is always of the en passage type, contains pleomorphic vesicles and establishes symmetrical contacts with dendrites. All boutons in the cats substantia nigra contain several large (700–1200 Å) dense core vesicles. Occasional axo-axonic contacts between type I and type III boutons are observed. Type I bouton is invariably presynaptic to the other.The findings are discussed in relation to some relevant problems.

The lateral reticular nucleus in the cat—I. An experimental anatomical study of its spinal and supraspinal afferent connections

The localization of the neurons from which the main ascending and descending projections to the lateral reticular nucleus originate has been studied in nine cats, using the retrograde axonal transport of horseradish peroxidase injected within that nucleus. The ascending spinoreticular neurons are widely distributed from the cervical to the sacral segments of the spinal cord. These neurons, which are of different sizes, are mainly located within Rexeds laminae VI, VII and VIII, but they spread both dorsally to laminae II–VI as well as ventrally to lamina IX. Labeled neurons of a size similar to motoneurons are particularly found within lamina IX, intermingled with the motoneurons. The spinal projection to the lateral reticular nucleus is crossed and uncrossed, with the ipsilateral spinoreticular neurons being located more dorsally within the grey matter of the spinal cord than the contralateral spinoreticular neurons. Moreover, while neurons with a crossed ascending projection are almost equally distributed along the whole rostro-caudal extension of the spinal cord, those with an uncrossed projection are predominantly located within the cervical segments of the spinal cord. Additional evidence indicates that the spinoreticular projection to the lateral reticular nucleus is somatotopically organized. In addition to the spinoreticular projection, the lateral reticular nucleus receives a crossed rubroreticular projection and a crossed fastigioreticular projection originating from the rostro-ventralmost part of the nucleus. A few neurons in the interposite nuclei also project to the lateral reticular nucleus.

Morphological changes in rat brain induced byl-cysteine injection in newborn animals

A single subcutaneous injection of L-cysteine (1.2 mg/g body weight) to rats 4 days after birth was followed by atrophy of the brain which was well developed 27--32 days after the injection. It was apparent that the lesioned animals could be divided into two groups (type 1 and 2) on account of the degree of brain atrophy. In type 1, which was observed in 80% of the animals, the body weight was unchanged, but the total brain weight was reduced by about 20%. The brain structures most affected were cerebral cortex, hippocampus and thalamus, each having a 30--40% reduction in wet weight. The atrophy of the posterior part of cortex was particularly pronounced in this type of lesion. In type 2 lesion, which appeared in 10% of the survivors, the atrophy was much more severe. There was a 50% reduction in wet weight of brain and in body weight. The most prominent finding was the atrophy of the whole cortex and the hippocampus which were reduced by 80 and 60% of wet weights respectively. In this type of lesion significant morphological changes were observed in several brain regions such as caudato-putamen, thalamus, pons, medulla oblongata, spinal cord and cerebellum.

Cerebellar projections to the nuclei ventralis lateralis and ventralis anterior thalami

SummaryAn experimental electron microscopical study has been made on the mode of termination of the cerebellothalamic projections in the cat. Supporting experimental light microscopical studies of silver impregnated sections following a large lesion of the cerebellar nuclei and light microscopical autoradiographic studies of the thalamus following injections of tritiated leucine in parts of the cerebellar nuclei, have been made as well. Following large lesions of the cerebellar nuclei, only the largest occuring type of bouton in the cats VL and VA (type LR bouton) degenerates. Following such lesions, type LR boutons undergo a filamentous hypertrophy before becoming electrondense. One degenerating LR bouton establishes complex synapses with the dendrites of both thalamocortical relay cells and interneurons. Not all type LR boutons in VL and VA degenerate following lesions of the cerebellar nuclei. Light microscopical autoradiographic studies as well as experimental electron microscopical investigations indicate that cerebellothalamic fibers end in clusters within VL and VA, and that the areas of termination lie more rostrally within these thalamic nuclei than has been inferred from experimental studies of silver impregnated sections following lesions of the cerebellar nuclei. The findings are discussed with respect to relevant morphological and physiological data.

Light and electron microscopical studies of the normal nuclei ventralis lateralis and ventralis anterior thalami in the cat

SummaryA light and electron microscopical investigation of the nucleus ventralis lateralis (VL) and nucleus ventralis anterior (VA) of the cats thalamus was made. Light microscopical examination of Golgi impregnated material revealed the existence of two types of cells based on differences in their dendritic arborization and branching of the axon. One of the cells is considered to be the thalamocortical relay cell, whereas the other is tentatively considered to be a Golgi type II neuron. Electron microscopical investigations of the two nuclei revealed the existence of a high number of profiles containing pleomorphic vesicles, and which have been identified as dendrites. Based on correlation with the Golgi material as well as on cytological features of the parent cell bodies, the dendrites containing vesicles are believed to belong to Golgi type II neurons. In addition to the vesicle-filled dendritic profiles, five different types of boutons have been identified. Two of these boutons, type LR and type SR, contain ovoid vesicles and establish asymmetrical synaptic contacts with dendrites of both types of neurons. Type F1, F2 and F3 boutons contain pleomorphic vesicles, but can be distinguished from dendritic profiles containing pleomorphic vesicles. Type F2 and F3 boutons establish symmetrical contacts with dendrites of both thalamocortical relay cells and Golgi type II neurons. Type F1 boutons establish symmetrical synaptic contact with the proximal dendrites or soma of the thalamocortical relay neurons only.Dendrites of both thalamocortical relay cells and Golgi type II neurons, as well as type LR, SR, F2 and F3 boutons, are engaged in glomeruli. Dendro-dendritic synapses between Golgi type II dendrites and relay cell dendrites are frequently seen, whereas no evidence of axo-axonic synapses have been found.Differences and similarities in the ultrastructural organization of VL and VA are described in some detail.

Cortical and pallidal projections to the nucleus ventralis lateralis thalami

SummaryThe nucleus ventralis lateralis (VL) and ventralis anterior (VA) thalami have been studied with the electron microscope following lesions of the cerebral cortex and of the nucleus entopeduncularis which represents the homologue of the medial pallidal segment in primates.It has been confirmed that VL receives a substantial number of afferents from the motor cortex, while cortical fibers to VA originate mainly rostral to the precruciate gyrus. Corticofugal fibers terminate in VL/VA as type SR boutons (Rinvik and Grofová, 1974a) and establish synapses with relay cell dendrites and with vesicle-containing dendrites.Four to five days following large lesions of the entopeduncular nucleus an electron-lucent form of degeneration occurs in one type of boutons in VL. These boutons are greatly swollen and vesicle-depleted, and contain altered mitochondria, an increased number of glycogen particles, irregular membrane structures and vacuoles. Some of the electron-lucent boutons progress into electron-dense forms at later survival times. Boutons showing these degenerative changes establish symmetrical synapses with relay cell dendrites and/or cell bodies. They do not synapse upon vesicle-containing dendrites and they are never engaged in the VL glomeruli. It is concluded that they belong to the type F1 boutons (Rinvik and Grofová, 1974a).Similar initial electron-lucent changes are seen in boutons in the nucleus centrum medianum (CM) ipsilateral to the entopeduncular lesions. No evidence was found for a projection from the entopeduncular nucleus to VA.The findings are discussed with regard to relevant morphological and physiological data in the literature. Particular attention is paid to the interaction at the cellular level in VL between afferents from the intracerebellar nuclei, motor cortex and globus pallidus.