Ipsilateral Pars Compacta Research Articles

Adrenalectomy counteracts the local modulation of astroglial fibroblast growth factor system without interfering with the pattern of 6-OHDA-induced dopamine degeneration in regions of the ventral midbrain

The present study investigated the effects of bilateral adrenalectomy (ADX) on the synthesis of basic fibroblast growth factor (bFGF, FGF-2) mRNA and on the expression of its FGF receptor subtype-2 (FGFR2) mRNA after a 6-hydroxydopamine (6-OHDA)-induced lesion of nigrostriatal dopamine system. In previous papers we have demonstrated that corticosterone increases FGF-2 immunoreactivity mainly in the astrocytes of the substantia nigra [Chadi, G., Rosen, L., Cintra, A., Tinner, B., Zoli, M., Pettersson, R.F., Fuxe, K., 1993b. Corticosterone increases FGF-2 (bFGF) immunoreactivity in the substantia nigra of the rat. Neuroreport 4, 783–786.] and that 6-OHDA injected in the ventral midbrain upregulates FGF-2 synthesis in reactive astrocytes in the ascending dopamine pathways [Chadi, G., Cao, Y., Pettersson, R.F., Fuxe, K., 1994. Temporal and spatial increase of astroglial basic fibroblast growth factor synthesis after 6-hydroxydopamine-induced degeneration of the nigrostriatal dopamine neurons. Neuroscience 61, 891–910.]. Rats were adrenalectomized and received a 6-OHDA stereotaxical injection in the ventral midbrain 2 days later. Seven days after the dopamine lesion, Western blot analysis showed a decreased level of tyrosine hydroxylase in the lesioned side of the midbrain, an event that was not altered by ADX or corticosterone replacement. Moreover, the degeneration of nigral dopamine neurons, which was confirmed by the disappearance of acidic FGF (FGF-1) mRNA and the decrement of tyrosine hydroxylase mRNA labeled nigral neurons, was not altered by ADX. The FGF-2 protein (23 kDa isoform but not 21 kDa fraction) levels increased in the lesioned side of the ventral midbrain. This elevation was counteracted by ADX, an effect that was fully reversed by corticosterone replacement. In situ hybridization revealed that ADX counteracted the elevated FGF-2 mRNA levels in putative glial cells of the ipsilateral pars compacta of the substantia nigra and in the ventral tegmental area. The ADX also counteracted the increased density and intensity of the astroglial FGF-2 immunoreactive profiles within the lesioned pars compacta of the substantia nigra and the ventral tegmental area as determined by stereology. The stereotaxical mechanical needle insertion triggered the expression of FGFR2 mRNA in putative glial cells, spreading to the entire ipsilateral ventral midbrain from the region of needle track, an occurrence that was partially reversed by ADX. In conclusion, bilateral ADX counteracted the increased astroglial FGF-2 synthesis in the dopamine regions of the ventral midbrain following a 6-OHDA-induced local lesion and interfered with FGF receptor regulation around injury. These findings give further evidence that adrenocortical hormones may regulate the astroglial FGF-2-mediated trophic mechanisms and wound repair events in the lesioned central nervous system.

Transient and Widespread Astroglial Activation in the Brain after a Striatal 6-Ohda-Induced Partial Lesion of the Nigrostriatal System

The authors have previously described astroglial activation in the ipsilateral nigrostriatal system and ventral tegmental area following small doses of 6-hydroxydopamine (6-OHDA) injected unilaterally in the striatum. This article further evaluated astroglial reactivity in several brain regions after striatal 6-OHDA-induced punctate lesion in the nigrostriatal pathway. Adult male Wistar rats received a unilatera1 stereotaxical injection of the 6-OHDA (8 μg/4 μl) in the neostriatum and sacrificed 1 or 3 weeks later. Control animals received only solvent. Immunohistochemistry was employed for visualization of the tyrosine hydroxylase (TH), marker for dopamine cells, and glial fibrillary acidic protein (GFAP), marker for astrocytes. TH immunoreactive terminals disappeared in the striatum close to the injection site and a disappearance of a small number of a defined population of dopamine cell bodies was observed in the ipsilateral pars compacta of the substantia nigra (SNc). No dopamine lesion was detected in the contralateral nigrostriatal pathway. Astroglial reaction was seen close to the lesion in the neostriatum and in the ipsilateral SNc of the 1 week 6-OHDA lesioned rats. Specific stereological tools employing point intercepts and rotator, revealed an increased presence of reactive astrocytes in many forebrain regions like frontal, parietal and piriform cortex, septum, neostriatum and SNc, bilaterally, and also corpus callosum after 1 week of 6-OHDA injection. The astroglial activation was characterized by increases in the size of the cell body and/or processes. Astrocytic reaction was found only in the ipsilateral nigrostriatal pathway by 3 weeks of 6-OHDA, a slight activation also remaining in the ipsilateral septum and piriform cortex. Astrocytic reaction was seen in the solvent-injected rats only in the neostriatum close to the needle track. The transient widespread astroglial reaction observed in many brain regions following a striatal injection of 6-OHDA may represent a global paracrine trophic response in the brain.

DOPAMINE CELL MORPHOLOGY AND GLIAL CELL HYPERTROPHY AND PROCESS BRANCHING IN THE NIGROSTRIATAL SYSTEM AFTER STRIATAL 6-OHDA ANALYZED BY SPECIFIC STEROLOGICAL TOOLS

Morphological changes in the dopamine neurons and glial cells of the rat midbrain ascending dopamine pathways were investigated after a partial lesion induced by unilateral striatal injection of a small dose of 6-hydroxydopamine (6-OHDA). Fourteen days after lesion, animals showed contralateral rotation induced by apomorphine injection. After behavioral analysis, rats were killed and their brains processed for the immunohistochemistry tyrosine hydroxylase (TH), a marker for dopamine cells, as well as glial fibrillary acidic protein (GFAP) and OX-42, markers for astrocyte and microglia, respectively. Stereological tools were employed in the quantifications. The volumes of the regions of the striatal TH immunoreactive disappearance, as well as the astroglial and microglial activation were several folds increased compared to control saline-injected rats. The optical disector detected decreases in the estimated total number of dopamine cells in the entire ipsilateral pars compacta of the substantia nigra (SNc) and ventral tegmental area (VTA) as well as in the estimated total number of varicosity profiles in the entire ipsilateral neostriatum. The stereological tool rotator showed no changes either in the mean or in the histogram distribution of the cytoplasmic volume of the nigral and VTA dopamine cells of 6-OHDA lesioned rats. Increases in the estimated total number of GFAP positive astrocytes were found in the entire neostriatum bilaterally as well as in the ipsilateral entire SNc and VTA of 6-OHDA lesioned rats. The estimated total number of OX-42 immunoreactive microglial profiles was elevated only in the ipsilateral entire neostriatum of the lesioned rats. The rotator detected cytoplasmic hypertrophy in the astrocytes, and also a shift to the right of the gaussian curves of the normal distribution of the logarithmic plotted values of the astroglial cell body volumes,of the neostriatum bilaterally as well as in the ipsilateral SNc and VTA of the striatal 6-OHDA injected rats. Cytoplasmic hypertrophy of microglia, and also a shift to the right of the gaussian curves of the values of microglia cell body volumes were seen only in the ipsilateral neostriatum; however, the point intercepts revealed an increased amount of microglial processes in the ipsilateral SNc and VTA of the lesioned rats. Specific stereological methods can be applied on detection of regionally different forms of cellular astroglial and microglial reaction after a partial lesion of dopamine pathway.

GLIAL bFGF AND S100 IMMUNOREACTIVITIES INCREASE IN ASCENDING DOPAMINE PATHWAYS FOLLOWING STRIATAL 6-OHDA-INDUCED PARTIAL LESION OF THE NIGROSTRIATAL SYSTEM: A STEROLOGICAL ANALYSIS

S100, a calcium-binding protein, and basic fibroblast growth factor (bFGF, FGF-2) are found predominantly in astrocytes in the central nervous system. Those molecules show trophic properties to neurons and are upregulated after brain lesions. The present study investigated the changes in the S100b and bFGF immunoreactivities after a partial lesion of the rat midbrain ascending dopamine pathways induced by intrastriatal injection of 6-hydroxydopamine (6-OHDA). Stereological method revealed increases in the estimated total number and density of bFGF immunoreactive astroglial profiles in the ipsilateral pars compacta of the substantia nigra (SNc) and ventral tegmental area (VTA). Increases in the counts of astroglial S100b immunoreactive profiles were found in the striatum, SNc, and VTA mainly ipsilateral but also in the contralateral nuclei. These results open up the possibility that interactions between astroglial S100b and bFGF may be relevant to paracrine events related to repair and maintenance of remaining dopamine neurons following striatal 6-OHDA induced partial lesion of ascending midbrain dopamine pathway.

Striatal injection of 6-hydroxydopamine induces retrograde degeneration and glial activation in the nigrostriatal pathway

PURPOSE: The effect of a highly selective 6-hydroxydopamine (6-OHDA)-induced lesion of the nigrostriatal system on the astroglial and microglial activation was analysed in adult Wistar rats after an unilateral striatal injection of the neurotoxin. METHODS: Male rats received an unilateral stereotaxical injection of the 6-OHDA in the left side of the neostriatum and were sacrificed 22 days later. Control animals received the injection of the solvent. The rotational behaviour was registered by a rotometer just before the sacrifice. Immunohistochemistry was employed for visualization of the tyrosine hydroxylase (TH) positive dopamine cells, glial fibrillary acidic protein (GFAP) immunolabeled astrocytes and OX42 immunoreactive microglia. Stereological method employing the optical disector was used to estimate the degree of the changes. RESULTS: The striatal injection of the 6-OHDA induced a massive disappearance (32% of control) of the TH immunoreactive terminals in a defined area within the striatum surrounding the injection site. A disappearance (54% of control) of dopamine cell bodies was observed in a small region of the ipsilateral pars compacta of the substantia nigra (SNc). The GFAP and OX42immunohistochemistry revealed astroglial and microglial reactions (increases in the number and size of the cells) in the ipsilateral neostriatum and SNc of the 6-OHDA injected rats. CONCLUSIONS: The striatal injection of 6-OHDA leads to retrograde degeneration as well as astroglial and microglial activation in the nigrostriatal dopamine pathway. Modulation of activated glial cells may be related to wound repair and to the trophic paracrine response in the lesioned nigrostriatal dopamine system.

Striatal Infarction in the Rat Causes a Transient Reduction of Tyrosine Hydroxylase Immunoreactivity in the Ipsilateral Substantia Nigra

Dopaminergic neurons of the substantia nigra pars compacta were examined in the rat brain following striatal infarction subsequent to transient focal cerebral ischemia. Rats had the middle cerebral artery occluded for 2 h or were sham-operated, and tyrosine hydroxylase immunoreactivity was evaluated by Western blot and immunohistochemistry at different times ranging from 1 to 60 days after ischemia. The number of tyrosine hydroxylase-immunoreactive cells in the substantia nigra pars compacta was counted under the light microscope and compared to that in the contralateral side and controls. No changes of tyrosine hydroxylase immunoreactivity were detected in the ipsilateral versus the contralateral substantia nigra of sham-operated rats or 1 day after ischemia. However, a statistically significant reduction of tyrosine hydroxylase-immunoreactive cells became apparent in the ipsilateral compared with the contralateral substantia nigra at 7 and 14 days after ischemia. This reduction showed a clear recovery at 30 days after ischemia, and no signs of difference between the ipsilateral and the contralateral side were apparent by 60 days. Therefore, the reduction of tyrosine hydroxylase immunoreactivity in the ipsilateral substantia nigra was only transiently seen from 1 to 2 weeks following ischemia. The observed loss of tyrosine hydroxylase was not accompanied by signs of cell death or gliosis in the ipsilateral pars compacta. The present results show a transitory reduction of tyrosine hydroxylase immunoreactivity in the ipsilateral substantia nigra pars compacta after focal ischemia and suggest that striatal infarction causes a transient deficit of dopaminergic function.

Gabaergic transmission and tyrosine hydroxylase expression in the nigral dopaminergic neurons: an in vivo study using a reversible ischemia model of rats.

The authors conducted an in vivo study, using a rat striatal ischemic model, of the effect of GABAergic transmission upon the dopamine synthesizing enzyme tyrosine hydroxylase in the neurons of the substantia nigra pars compacta. Two hours transient middle cerebral artery occlusion produced massive striatal ischemic damage resulting in a marked decrease of GABAergic projection to the ipsilateral substantia nigra. Histological examinations were conducted in rats killed at three, seven, 15, 30 and 94 days after ischemia. The immunoreactivity for tyrosine hydroxylase in the ipsilateral pars compacta was unaltered up to three days after the ischemic insult, but it was markedly decreased at seven days post-ischemia. At this stage, the number of neurons positive for tyrosine hydroxylase was significantly decreased in the ipsilateral pars compacta, whereas there was no significant reduction in the number of pars compacta neurons containing Nissl substance. By 30 days post-ischemia, the tyrosine hydroxylase-positive cell number in the ipsilateral pars compacta appeared to be equivalent to that of the contralateral side. It was also noted that continuous intraventricular administration of a GABAA receptor agonist muscimol, initiated from 24 h post-ischemia, effectively prevented the transient reduction of immunoreactivity for tyrosine hydroxylase in the ipsilateral pars compacta at seven and 15 days after ischemic insult. The present study revealed that the striatal ischemic lesion induced a transient down-regulation of tyrosine hydroxylase synthesis in the pars compacta neurons, which could be prevented by administration of GABA agonist, suggesting that GABAergic transmission greatly affects dopamine metabolism in these cells.

Temporal and spatial increase of astroglial basic fibroblast growth factor synthesis after 6-hydroxydopamine-induced degeneration of the nigrostriatal dopamine neurons

The present study investigates the temporal and spatial changes of the cellular expression of basic fibroblast growth factor messenger RNA and immunoreactivity after a 6-hydroxydopamine-induced lesion in the nigrostriatal dopamine system. In situ hybridization revealed a sustained (from 4 h to two weeks) and strong (300–400% of control, at the peak intervals) increase of basic fibroblast growth factor messenger RNA in the pars compacta of the substantia nigra and the ventral tegmental area ipsilateral to the lesion. A short-lasting increase of basic fibroblast growth factor messenger RNA was observed in the ipsilateral pars reticulata of the substantia nigra (from 4–24 h, 300% of control) and neostriatum (24 h, 180% of control) as well as in the ipsilateral and contralateral hippocampus and neocortex (by 4 h, 200% of control). Brightfield microscopy showed an increased number of putative glial cells expressing the basic fibroblast growth factor messenger RNA signal. Basic fibroblast growth factor immunohistochemistry revealed on control brains the protein in the nuclei of glial cells throughout the forebrain and the midbrain and in the nuclei of neurons of the layer II of the retrosplenial granular cortex, the CA2 region of the hippocampus and the fasciola cinereum as well as in the nuclei of ependymal cells. The injection of 6-hydroxydopamine increased basic fibroblast growth factor immunoreactivity in the nuclei of astrocytes only within the ipsilateral substantia nigra and ventral tegmental area. By 2 h after the drug injection, the density of glial basic fibroblast growth factor-immunoreactive profiles was increased in the pars compacta of the substantia nigra and the ventral tegmental area. The density, size and intensity of the astroglial basic fibroblast growth factor immunoreactive nuclei were increased in the entire substantia nigra and the ventral tegmental area at 72 h, and peaked one week after the 6-hydroxydopamine injection. The saline injection promoted a time-dependent increase in the density of the glial basic fibroblast growth factor immunoreactivity but only in the ipsilateral pars compacta of the substantia nigra. In conclusion, the dopamine cell degeneration may give rise to extracellular signals activating the surrounding astroglia, leading to a sustained increased synthesis of astroglial basic fibroblast growth factor, which may exert neuroprotective action and increase repair on the nigrostriatal dopamine system.

Bilateral morphological changes in the substantia nigra of the rat following unilateral damage of the striatum

The effects of damage of the striatum and globus pallidus of one side on the size of cells in the pars reticulata and pars compacta of the substantia nigra on both sides and in the contralateral globus pallidus have been examined. Cellular cross-sectional areas have been compared with those for neurons in the same nuclei in normal age and sex matched littermate control animals. One week after removal of the left striatum and globus pallidus and overlying cortex, the cells in the ipsilateral pars compacta are significantly shrunken (15%). This decrease in size gets progressively more marked with longer survival times reaching 50% 112 days after operation, the longest survival time examined. The shrinkage is accompanied by marked cell loss. Neurons in the contralateral pars compacta show an initial significant hypertrophy of their cell bodies (20%) in the first week after the operation, and later show a shrinkage of 20% to 35 days. The degree of this contralateral shrinkage gradually declines to 12% at 112 days. The changes in the pars compacta are accompanied by a significant enlargement (33%) of the cells in the pars reticulata of the substantia nigra on the side of the damage. This hypertrophy is present by 35 days after operation and persists at least until 112 days. Similar hypertrophy occurs in the ipsilateral globus pallidus in the one case where this could be examined. There are no significant changes in the contralateral pars reticulata, but there is significant enlargement (23%) of the neurons in the contralateral globus pallidus.

Localization of nigrostriatal, nigrothalamic and nigrotectal neurons in ventricular coordinates in macaques

The topography of the substantia nigra and its subdivisions was first analysed in macaques by using a topographic technique based on ventricular landmarks. This study shows the stability of the contours of the substantia nigra and its subdivisions in various species of macaques. The anteroposterior sequence of four subdivisions was standardized by defining eight verticotransverse levels, regularly interspaced and systematically used for each experimental case. Neurons of the substantia nigra sending axons to the striatum, thalamus and superior colliculus were identified by the technique of retrograde transport of horseradish peroxidase. The nigrostriatal labeled neurons were essentially located in the ipsilateral pars compacta but also scattered dorsally in the pars mixta and ventrally in the pars reticulata. In addition, the existence of a crossed nigrostriatal pathway was demonstrated in monkeys. Nigrothalamic labeled neurons were found in the whole pars reticulata at rostral levels and only in the medial part at more caudal levels. Comparatively, nigrotectal labeled neurons were also found in the whole pars reticulata at rostral levels, but caudally, they were confined to the lateral part of the pars reticulata and the pars lateralis. It thus appears that these three nigral components may overlap at some levels of the substantia nigra. This is discussed in relation to the existence of branched axons already documented. However, the present results underline the strong tendency of the nigrotectal neurons to be segregated from the nigrothalamic ones and to be laterally located in monkeys. In addition, two nigrotectal components have been identified on the basis of their topography and their somata size: one with large somata located in the pars lateralis, probably specific to primates, and the other with smaller somata located in the pars reticulata. These two components may indicate the existence of two different functional systems.

Cells of origin of subcortical afferents to the caudate nucleus: A horseradish peroxidase study in the cat

The horseradish peroxidase tracing method was used to investigate the sources of subcortical input to the caudate nucleus in the cat. Of considerable interest was the finding, not previously described, that labeled neurons were present outside of the intralaminar complex, in the ventral anterior and mediodorsal thalamic nuclei. Another new finding was that retrogradely labeled cells were present within the basolateral amygdaloid nucleus, following the caudate injections. In addition, neurons containing enzyme granules were found within all of the intralaminar thalamic nuclei, most prominently in the central lateral and centromedian nuclei. Also, HRP-positive cells were strikingly apparent within the ipsilateral pars compacta of the substantia nigra, and a few labeled cells were found in the ipsilateral pars reticularis, and the contralateral pars compacta of the substantia nigra. Cells containing peroxidase granules were also found in the ventral tegmental area and the retrorubral area. The most caudal projection to the caudate nucleus in the cat was demonstrated by labeled neurons found bilaterally in the dorsal nucleus of the mesencephalic raphe.