Search

Rats that had been walking on a rotating drum for a period of 75 min, demonstrated expression of the immediate early gene c-Fos in specific areas of the inferior olive and cerebellar nuclei. Non-walking control rats did not show consistent Fos-like immunoreactive labelling in these nuclei. Fos-like immunoreactive olivary neurons were consistently found in the caudolateral parts of the dorsal accessory olive and in its dorsal fold, and within specific areas of the medial accessory and principal olives. In the cerebellar nuclei, immunoreactive cells were found in the medial part of the anterior interposed nucleus, in the interstitial cell groups, and within specific parts of the medial, posterior interposed, and lateral nuclei. The lateral vestibular nucleus as well as the ventral part of the magnocellular red nucleus also displayed positive neurons. Most of these positive areas are known (1) to be related to the spinal cord and (2) to be anatomically linked. These results are in line with a presumptive role of the olivo-cerebellar system in ongoing movement control.

Radioligand binding and behavioral studies were conducted to determine whether a relationship existed between the motor effects produced by (+)-pentazocine and its binding to sigma sites. Scatchard analyses revealed decreased [3H](+)-pentazocine binding in middle aged rats (5-6 months old) compared to young adult rats (2-3 months old). However, there was no difference between the extent of circling behavior or dystonia produced by microinjection of (+)-pentazocine into the substantia nigra or red nucleus in the older animals compared to the young adult rats. There was also a significant decrease in [3H](+)-pentazocine binding in rats chronically treated with haloperidol. Again, however, despite the reduction in [3H](+)-pentazocine binding, there was no difference between the extent of dystonia produced by unilateral intrarubral microinjection of (+)-pentazocine into animals chronically treated with haloperidol vs. saline. The postural changes produced by (+)-pentazocine could not be attenuated with coadministration of the putative sigma receptor antagonist BD1047 (N-[2-(3,4-dichlorophenyl)ethyl]-N-methyl-2-(dimethylamino) ethylamine), or the opiate receptor antagonist naloxone. However, the (+)-opiate, (+)-nordihydrocodeinone, partially attenuated the postural effects of (+)-pentazocine, despite its very low affinity for sigma 1, sigma 2, or opiate receptors. Taken together with previous studies, the results suggest that [3H](+)-pentazocine is a potent and selective probe for sigma 1 binding sites, but the in vivo effects of (+)-pentazocine cannot be fully attributed to actions through these sites. Some of the in vivo effects of (+)-pentazocine appear to involve other binding sites that are not detected under the conditions normally used in in vitro assays.

This work tests the hypothesis that deafferentation caused by experimental brain lesions leads to the appearance of perikaryal argyrophilia and the accumulation of phosphorylated cytoskeletal proteins which are potential precursors for neurofibrillary tangle formation. Destructive lesions of the ventral tegmental area, the septum of the medial forebrain and the entorhinal cortex, when combined with systemic administration of a D-1 dopamine receptor antagonist, produced transsynaptic changes in neurons of the hippocampal formation in the midbrain. Abnormally phosphorylated neurofilament protein was demonstrated immunohistochemically in the cytoplasm of mesencephalic pyramidal neurons, particularly in the red nucleus. These same neurons also developed cytoplasmic argyrophilia in Bielschowsky histologic preparations. Although distinct neurofibrillary tangles were not produced with this paradigm, both the protein immunoreactivity and argyrophilia were arranged in cytoplasmic linear arrays. The structural changes induced in this experimental model may be an important preliminary stage for neurofibrillary tangle formation.

The cellular localization of calbindin D-28k (CB) and parvalbumin (PV) in the red nucleus of the rat was studied by means of double-immunohistochemical techniques applied to single sections. Neurons displaying immunoreactivity for either CB or PV were found throughout the rostrocaudal extent of the red nucleus, but PV neurons predominate in the rostral two-thirds and CB neurons in the caudal two-thirds of the nucleus. Likewise, there was a clear but not absolute segregation of the two types of neurons along the dorsoventral axis; PV and CB neurons were largely confined to the dorsolateral and ventromedial sectors of the nucleus, respectively. Most CB neurons were large (> 30 microns), whereas large and medium-sized (15-30 microns) PV neurons were equally abundant. Additionally, some large and medium-sized neurons displaying immunoreactivity for both PV and CB were encountered in the ventromedial sector of the red nucleus. The present study reveals that, in contrast to previous beliefs, the red nucleus is composed of a neuronal population that is chemically highly heterogeneous.

The presence of cholinergic input to the red nucleus (RN) in the cat is demonstrated by choline acetyltransferase (ChAT) staining; however, it is unclear what effect this cholinergic input has on neurons of the RN. Further, the presence of cholinergic neurons in the rat RN has been the subject of controversy. The present study examined the effects of intravenous injections of S(-)-nicotine tartrate (62.5-250 micrograms/kg) on the firing rate of rubral neurons. Dose-dependent increases in firing rates were observed which were blocked by pre-treatment with the nicotinic antagonist, mecamylamine hydrochloride. Smaller consistent increases were found after pretreatment with 62.5 micrograms/kg or 125 micrograms/kg doses of nicotine than were observed following the initial administration, suggesting a desensitizing response typical of nicotinic receptors.

Neuromelanin is a poorly understood pigment that accumulates in catecholaminergic neurons during normal aging. Electron paramagnetic resonance spectroscopy, an especially effective technique for investigating melanins, is used in the present study to show unambiguously that neuromelanin is a melanin; however, it is not well modeled by synthetic dopamine melanin and thus is an atypical melanin. Some of the unusual features of neuromelanin can be explained by postulating two distinct sources for its free radicals, the dominant one possibly derived from a precursor containing sulfur. Examination of human substantia nigra by electron paramagnetic resonance spectroscopy during the purification of neuromelanin also demonstrates, contrary to some other studies, that a portion of the paramagnetic metal ions in this tissue are bound to the pigment in situ. Combined with previous histochemical data, these observations have implications for the mechanism through which neuromelanin accumulates in vivo and are consistent with its having a cytoprotective function under normal conditions, but a cytotoxic role at advanced ages and in patients with Parkinson's disease. Other results of this study show that homogenizing tissues during the purification of any natural pigment may cause contamination of the pigment by extraneous metal ions and that subsequent incubation in hot acid, though most effective in removing metal ions and hydrolyzing proteins, leads to degradation of melanin. A purification procedure using incubation in acid at room temperature, however, is well suited for identifying and characterizing unknown natural pigments by electron paramagnetic resonance spectroscopy.

Many immediate early genes are rapidly and transiently expressed in the central nervous system following a variety of stimuli. Damage to the axons of peripheral and certain central neurons has been shown to result in a long-term increase in expression of c-Jun in the parent cell bodies. In the peripheral nervous system this increased expression of c-Jun protein and mRNA develops over 24 h following sciatic nerve section and is maintained if the damaged nerve is ligated, but returns to basal levels if the peripheral nerve is allowed to regenerate. Here, we report on the response of rubrospinal neurons to spinal cord hemisection at levels C3 and T10. c-Jun expression was first seen at 12 h post-lesion in a limited number of rubral neurons. The number of positively stained neurons increased up to 10 days post-lesion and then declined over the following weeks. By 7 weeks post-lesion there was still evidence of c-Jun immunoreactivity in both large and other clearly atrophic rubrospinal neurons. c-Fos immunoreactivity was seen only at 12-48 h in a small number of rubrospinal neurons. Evidence from retrograde tracing experiments following fluorogold application to the hemisected cord suggested that all c-Jun-positive neurons projected into the spinal cord. No c-Jun response was seen following a lesion at T10.(ABSTRACT TRUNCATED AT 250 WORDS)

Acrylamide, an occupational neurotoxicant, reduced MAP1 and MAP2 distribution in different regions of rat brain. Different components of the extrapyramidal system (caudate-putamen, globus pallidus, substantia nigra and red nucleus) revealed differential distribution of MAP1 and MAP2 in acrylamide-treated animals. Rats were treated with acrylamide (estimated mean dose: 15 mg/kg/day) for 2 weeks and MAP1 and MAP2 were localized according to Sternberger's peroxidase-anti-peroxidase technique. MAP1 labelled neuronal perikarya and dendrites almost with a similar intensity, but MAP2 immunostaining was more intense in dendrites than neuronal perikarya. Acrylamide caused a near-total loss of MAP1 and MAP2 immunoreactivity in caudate-putamen. Other components of the extrapyramidal system were relatively less affected by acrylamide. These results indicate that caudate-putamen is more susceptible to the action of acrylamide than other components of the extrapyramidal system studied. The depletion of MAP1 and MAP2 immunoreactivity by acrylamide appears to be an early biochemical event preceding peripheral neuropathy. The loss of MAPs immunoreactivity occurs first in dendrites and proceeds toward the perikarya. This study indicates that acrylamide not only causes axonal damage but may also induce dendritic degeneration.

Iron, an essential element for basic cellular metabolism, regularly accumulates in certain brain areas in normal subjects and in patients with certain diseases. Magnetic resonance imaging can depict iron deposition, offering a singular opportunity to correlate the regional iron content with the functional status of the human brain in vivo. We studied the relationship between age and the iron-related signal loss on T2-weighted images in basal ganglia, and observed a strongly significant signal decrease in the globus pallidus at the age of brain development (first two decades of life), but we found no such decrease in later years. Moreover, in healthy adults, subject-to-subject variability was relevant in changes due to iron deposition in magnetic resonance imaging. We found increased signal loss to be associated with poor performance on motor and specific cognitive tasks, suggesting that these image changes can provide functional information with respect to the brain in normal subjects.

Inhibitory regulation of dopamine neurons is mediated by dopamine autoreceptor and gamma-aminobutyric acidB receptor opening of potassium channels. Increased potassium conductance by either receptor is G protein dependent. To evaluate the role of G proteins in vivo, pertussis toxin (PTX) was microinjected into the A10 dopamine region and changes in dopamine metabolism and synthesis measured. PTX produced an elevation in dopamine metabolism and synthesis in the A10 region and nucleus accumbens for up to 4 days after injection. By day 7 the levels of the dopamine precursor and metabolites had returned to normal. A less consistent increase was also measured in the A9 dopamine region and the prefrontal cortex. Although dopamine synthesis and metabolism had returned to normal by day 7, the in vitro ADP-ribosylation of G proteins in the A10 region by PTX remained depressed by approximately 50% from day 1 to day 14 after administration, returning to normal by day 30. The data suggest that in vivo ribosylation of G proteins may lead to a short-term attenuation of the tonic inhibitory control of dopamine neurons, which can be compensated for by PTX-insensitive mechanisms.

Neurons confined within the mammalian CNS usually do not regenerate after axonal injury, while axonal regeneration is the rule in the PNS. It has been hypothesized that this may be related to differences in the microenvironment of the PNS versus CNS and to differences in the neuronal response to injury. In order to test the latter hypothesis, we compared changes in gene expression after axotomy in two populations of neurons: rat facial motoneurons and rat rubrospinal neurons. In situ hybridization with cDNA probes for the medium and light neurofilament protein revealed a reduced mRNA content in both facial and rubrospinal neurons at all times investigated (i.e., 1, 2, and 3 weeks after axotomy). On the other hand, mRNAs for actin and tubulin were increased in both neuronal populations during the first week after axotomy. While this increase was sustained in facial motoneurons for several weeks, total tubulin mRNA and actin mRNA were decreased in rubrospinal neurons at 2 and 3 weeks after axotomy, coincident with their atrophy. The developmentally regulated T alpha 1 tubulin mRNA, which was previously shown to be reexpressed in facial motoneurons after axotomy, was elevated severalfold in axotomized rubrospinal neurons, and increased levels persisted in some rubrospinal neurons as late as 7 weeks after axotomy. Similarly, the developmentally regulated GAP-43 mRNA increased in both axotomized facial and rubrospinal neurons, and increased levels were sustained in some axotomized rubrospinal neurons for at least 7 weeks. The response of rubrospinal neurons to axotomy in the cervical spinal cord is, in the first week, qualitatively similar to the response of facial motoneurons. However, by 2 weeks after axotomy there is a generalized reduction in mRNA levels for all three cytoskeletal proteins that is associated with neuronal atrophy. During this period, mRNA levels for the two specific markers of the growth state, T alpha 1 tubulin and GAP-43, remain elevated. Thus, axotomy of rubrospinal neurons appears to set in motion two independent events. First, an axotomy signal initiates a cell-body reaction similar to that of PNS neurons, including increased mRNA levels for T alpha 1 tubulin and GAP-43. Later, a generalized cellular atrophy and decrease in mRNA levels occur without reversing the specific responses of T alpha 1 and GAP-43 to axotomy. We conclude that the failure of rubrospinal neurons to regenerate is not due to a failure to initiate gene-expression changes characteristic of regenerating peripheral neurons.(ABSTRACT TRUNCATED AT 400 WORDS)

Hybridization signals indicating mRNAs encoding the precursor of calcitonin gene-related peptide (CGRP) and CGRP immunoreactivity were detected on parallel sections containing brainstem motor nuclei using in situ hybridization histochemistry and immunohistochemistry. In untreated and saline-injected rats the motoneurons in the hypoglossal, facial motor nuclei and in the ambiguus nucleus showed weak to moderate hybridization signals. In these motoneurons CGRP immunoreactivity was restricted to the Nissl bodies of the perikarya. Twenty-four and 42 hours after intracerebroventricular colchicine injection the intensity of both the hybridization signal and the immunoreaction product increased. The distribution of CGRP immunoreactivity changed from discrete perikaryal localization to diffuse reaction in the perikarya and along the proximal dendritic tree. Motoneurons in the rest of the brainstem motor nuclei (VIth, Vth, IVth and IIIrd) of untreated and saline-injected rats showed neither hybridization signal nor CGRP immunoreactivity. After intracerebroventricular injection of colchicine these motoneurons showed both hybridization signal and CGRP immunoreactivity. In all nuclei the size of motoneurons decreased and their Nissl structure changed to an amorphous basophilic mass following colchicine treatment.

Three (+)-benzomorphans that bind to sigma receptors produced dystonia in a dose-related manner when microinjected into the red nucleus of rats. Two lines of evidence suggest that these effects were related to the sigma-binding properties of the compounds. First, the behavioral potency of the (+)-benzomorphans and other active sigma compounds correlated highly with their affinities for [3H]1,3-di-o-tolylguanidine-labelled sigma receptors in the rat brain (r = .94). Second, similar intrarubral injections of non-sigma ligands were without effect: various vehicles, a structurally related (+)-opiate with no affinity for sigma receptors, and selective dopaminergic and serotonergic compounds failed to significantly alter the normal posture of rats. The only ligand in this study that binds with high affinity to sigma receptors, but failed to elicit torsional head movements was (+)-[3-(3-hydroxyphenyl)-N-(1-propyl)piperidine] [(+)-3PPP], a ligand with mixed activity at sigma and dopamine receptors. Since (+)-3PPP failed to produce an effect on its own and also failed to attenuate the dystonia produced by another sigma ligand (DTG), it may interact with a non-sigma mechanism or with a different sigma receptor type from the other compounds.

Dopamine (DA) and homovanillic acid (HVA) concentrations were measured in subregions of substantia nigra, ventral tegmental area and retrorubral field in vervet monkeys 1 to 2 months after treatment with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Identical MPTP treatment regimens produced animals with different degrees of parkinsonism. In asymptomatic monkeys, changes in DA and HVA concentrations in the midbrain DA regions were relatively small and involved central substantia nigra and dorsomedial ventral tegmental area. In contrast, changes in symptomatic monkeys were more severe and widespread, significantly affecting all examined subregions of substantia nigra (greater than 75% DA depletion), both dorsomedial and ventromedial ventral tegmental area and lateral, but not medial, retrorubral field. The data indicate that DA neurons in subregions of substantia nigra, ventral tegmental area and retrorubral field are not equally susceptible to MPTP toxicity. The pattern of MPTP-induced DA and HVA losses in the vervet monkey mesostriatal dopaminergic system may resemble postencephalitic Parkinson's disease more closely than idiopathic Parkinson's disease.

The release of endogenous gamma-aminobutyric acid (GABA) from the cat red nucleus was studied in a slice preparation. Potassium-induced depolarization caused a release of GABA in a calcium-dependent manner. The release of GABA was also induced by veratrine and blocked by tetrodotoxin. The demonstration of release satisfies one of the important criteria for identification of GABA as the inhibitory neurotransmitter.

The distribution of presumed glutamatergic projections from sensorimotor cortical areas to the red nucleus (RN), the substantia nigra (SN), the ventrolateral thalamic complex, and the caudate nucleus (CN) was investigated in the cat. For this purpose, the changes in the sodium-dependent high-affinity glutamate uptake (HAGU) rate were measured in homogenates of tissue microdissected from various parts of these subcortical structures after chronic surgical ablation of sensorimotor cortical areas. After 8 to 10 days survival, significant reductions in HAGU activity were noted in the structures studied on the side ipsilateral to the operated cortex. Within each structure, various quantitative or qualitative changes were observed. Higher decreases in HAGU activity were found in the caudal part of the RN, the ventrolateral thalamic nucleus, and the dorsolateral part of the CN than in the other parts of these structures. The lateral part of the SN showed a large decrease in HAGU rate and its medial part a small but significant increase. Referring to the anatomic data concerning the organization of cortical projections from sensorimotor areas to the structures studied, it was shown that our results support the view that glutamate could act as a neurotransmitter along various corticosubcortical pathways.

We have shown the presence of a specific binding of 5 hydroxytryptophan in the red nucleus and oculomotor nucleus of the Rat brain. The analysis of the saturation curve carried out on a membrane preparation suggests the existence of a high affinity binding site (Kd = 1 nM).

One-sided lateral funiculotomy at the C-2 segment induced axon reaction in the contralateral red nucleus of adult cats. Two to 60 days postoperatively the animals were sacrificed and the mesencephalon was fixed in ethanol-acetic acid, 3:1. Ten micrometer paraffin sections including both red nuclei were stained for RNA with azure B after incubation in DNAse. Cytophotometric measurements of RNA content of neurons from the caudal 600-1000 micrometer of each red nucleus were made with a Zeiss Cytoscan system using an automatic scanning stage. In contrast to the heightened RNA synthesis that has been reported for axotomized peripheral (extrinsic) neurons, the axotomized central (intrinsic) neurons of the red nucleus showed no evidence of accumulation of cytoplasmic or nucleolar RNA. Rather depletion of cellular RNA occurred. Further indication of the regressive nature of rubral axon reaction derived from morphometric measurements that showed cytoplasmic, nuclear and nucleolar atrophy of the neurons of the red nucleus contralateral to operation with the exception of a possible transient cytoplasmic enlargement 9 days postoperatively. From the cytophotometric and morphometric data here reported we are led to suggest that the frequently observed failure of axonal repair in mammalian CNS results from the innately regressive nature of the axon reaction of many mammalian central neurons.

The distribution of amino acids in two Friedreich's ataxia brains is compared to 4 control brains. Glutamate and GABA were decreased in the cerebellar hemispheres and/or in the vermis. Taurine concentrations were uniformly elevated throughout the areas studied. Taurine/glutamate and taurine/GABA ratios were consistently elevated in Friedreich's ataxia brains, whereas glutamate/glutamine ratios were consistently decreased (with normal glutamine concentrations).

Some basic methodological issues concerning the use of the fluorescent tracers Fast blue (FB) and Diamidino yellow (DY) were studied using the projections of the red nucleus to the nucleus interpositus anterior (NIA) of the cerebellum of the cat. On standard Nissl-stained sections, it was possible to delineate 4 distinct zones at the FB and DY injection sites. Correlative studies of injection sites in the NIA and retrograde labeling of cell bodies in the contralateral red nucleus showed that effective uptake occurred only from the zone mechanically damaged by the injection needle (termed zone 0). The tracer remains in this zone during the post-injection survival. The limited uptake area for both tracers is a valuable feature for studies of restricted neuronal projections. However, the tracers are not suitable for use in quantitative studies, especially those concerning axonal collateralization. Perfusion with water-soluble fixatives did not alter the cellular storage site. In double-labeling experiments using horseradish peroxidase and DY, the HRP histochemistry induced an important "washing out" of DY and consequently, an underestimation of the number of labeled neurons.

Horseradish peroxidase (HRP) was applied to the precruciate cortex of the cat. The peroxidase was either applied directly to the surface or injected by glass micropipettes in single or multiple injections. Retrograde as well orthograde transport was observed by light and electron microscopy in different areas: in the thalamus: the nucleus ventralis lateralis and the nucleus centrum medianum, in the red nucleus and in the spinal cord. The distribution of HRP within the nervous elements of the nuclei: axon terminals and/or perikarya and dendrites, was observed by light and electron microscopy. HRP was present in dendrites and perikarya, indicating retrograde transport, in experiments where damage to the injected nervous tissue could be suspected. However HRP appeared restricted to the nerve endings, indicating orthograde transport, when the damage seemed to be minimal. This effect of injury on retrograde transport of HRP was further confirmed by experiments using bilateral HRP injections and unilateral lesions of the spinal cord to label rubro-spinal cells. Thus retrograde transport of HRP could be induced by lesion in systems where it does not occur normally. The normal intra-axonal transport of HRP appears to be from the perikaryon to the nerve endings. This orthograde transport can be seen under the light microscope and can be confused with faint retrograde labeling.

Disorders of neurotransmitter balance are observed in Parkinson's disease, pharmacotoxic psychosis and depression. The dopamine-serotonin ratio is reduced to about 20% in Parkinson and pharmacotoxic patients in the caudate nucleus and in the substantia nigra. The serotonin content in these brain areas is lowered only to about 50% in comparison to that of the control, whereas the dopamine level is reduced to 85% in Parkinson patients. This dopamine deficiency has been substituted by exogenous supply of L-dopa in combination with decarboxylase and monoaminooxydase inhibitors. First evidence is presented that L-dopa can be replaced, at least partially, by iron in form of a ferriascorbate complex. This iron compound improves the symptoms of Parkinson's disease to almost the same extent as L-dopa.

The effects of local postlearning microinjections of arginine-vasopressin (AVP) and oxytocin (OXT) on one-trial learning passive avoidance behavior and the influence of AVP on alpha-MPT-induced disappearance of norepinephrine (NE) and dopamine (DA) in discrete brain regions have been studied in the rat. OXT injected bilaterally in the hippocampal dentate gyrus (25-25 pg) or in the midbrain dorsal raphe nucleus (50 pg) significantly attenuated passive avoidance behavior. Facilitation of passive avoidance behavior was observed when the peptide was injected into the dorsal septal nucleus. AVP facilitated passive avoidance behavior when administered into the hippocampal dentate gyrus, dorsal raphe nucleus or dorsal septal nucleus. Injection of either neuropeptides into the central amygdaloid nucleus appeared to be ineffective. One week after the behavioral experiments a repeated injection of AVP into the hippocampal dentate gyrus increased the disappearance of NE in the dentate gyrus and in the nucleus ruber. An injection into the dorsal septal nuclei decreased the NE disappearance in the dorsal septal nucleus itself and increased it in the nucleus ruber. Injection in the dorsal raphe nucleus led to an increase in the disappearance of DA in the locus coeruleus and in the nucleus ruber. It is concluded that memory consolidation can be oppositely influenced by local application of minute amounts of either OXT or AVP into certain limbic-midbrain structures, suggesting an involvement of these brain regions in the memory effects of these peptides. Modulation of catecholamine turnover in specific brain areas after AVP administration may be related to this behavioral effect.

High affinity glutamate uptake (HAGU) was measured within the red nucleus (RN) and the ventrolateral thalamic area in intact adult cats and in animals which had undergone a large hemicerebellectomy 8 to 21 days before. In the side contralateral to the lesion, results show two types of changes in HAGU: 1. In the caudal parts of the RN and the ventrolateral thalamic nucleus (VL), a strong HAGU decrease was demonstrated suggesting some cerebellorubral and cerebellothalamic fibres use glutamate (Glu) as their neurotransmitter. 2. In the rostral parts of the RN and the VL, an increase in HAGU was detected. This increase was particularly large at thalamic level, which led us to perform a kinetic analysis of the uptake system. Results show that the increase observed in HAGU is related in the thalamic area to an increased affinity of the transport sites for Glu. The mechanism of the HAGU increase measured in the rostral VL after cerebellectomy was further investigated in the presence of acetylcholine (ACh) which we have previously shown to be possibly involved in the neurotransmission of some cerebellothalamic and cerebellorubral fibres. ACh was shown to exert an inhibitory effect on HAGU in the control situation. Decrease in affinity of the transport sites for Glu induced by ACh was more pronounced when HAGU was enhanced as a consequence of the cerebellar lesion. We hypothesized that the cerebellectomy enhances the activity of nerve terminals which take up Glu in the VL and that we have shown to be mainly related to corticothalamic neurons. The basic mechanism involved in this activation could be the withdrawal of presynaptic inhibitory controls on corticothalamic fibres due to the removal of the putative cholinergic cerebellar input. This hypothesis was extended to the RN where previous electrophysiological and anatomical studies have suggested that the cerebellar lesion induces a sprouting of corticorubral nerve terminals. The increase in HAGU in response to the cerebellar lesion could constitute an adaptive mechanism by which the CNS may compensate for the loss of the excitatory cerebellar input to the RN and thalamic neurons by increasing the corticofugal transmission.

In this study, we have tested the possible use of protein-gold complexes as neuronal markers for long-term tracing studies in rat. The tracer we have used consisted of colloidal gold particles coupled to wheat-germ agglutinin apohorseradish peroxidase conjugate (WGA-apoHRP). The neuronal labeling was studied for survival periods of up to nineteen months following injection in the central nervous system. Maximal visualization of the gold particles was achieved through gold silver intensification. The tracer could be detected throughout the entire range of periods considered. The injection site consisted of a dense black core and retrogradely labeled cells were characterized by round black granules over the cell body. The retrogradely labeled cells were cytochemically characterized by demonstrating their transmitter content. Thus protein-gold complexes may be used as long-term neuronal markers compatible with the persistance of the vital functions of the labeled cells.

High affinity glutamate uptake into corticofugal fiber terminals was measured in the ventrolateral thalamus and red nucleus at varying time intervals after lesions were made by kainic acid in the contralateral interpositus nucleus of the cerebellum in rats. Under similar conditions the density of cortical fiber terminals was estimated using the Fink-Heimer impregnation technique. 1. Glutamate uptake steadily increased in the ventrolateral thalamus up to 60 days after lesions in the contralateral cerebellum. 2. Similar changes were noted in the red nucleus. 3. The changes were dependent on the integrity of corticofugal fibers to the thalamus and red nucleus. 4. No changes in uptake of gamma-aminobutyric acid were noted. 5. Saturation curves for glutamate uptake suggested a change in the maximal number of transport sites. 6. Fink-Heimer degeneration studies showed an increase in cortical terminals in the ipsilateral ventrolateral thalamus and in both rostral and caudal regions of the red nucleus following lesions in the contralateral interpositus nucleus. The data are consistent with an increase in the number of cortical fiber terminals in reaction to loss of cerebellar input to the ventrolateral thalamus and red nucleus. This study correlates anatomical and biochemical evidence for collateral sprouting in a model based on electrophysiologic data in the red nucleus and extends the model to include the thalamus.

The mesencephalic dopamine (DA) cell system was examined in mice homozygous and heterozygous for the weaver (wv) gene and in wild-type controls to estimate the extent of cell losses associated with the genetically determined central DA deficiency observed in weaver homozygotes. Animals of the three genotypes (+/+, wv/+, wv/wv) were studied at postnatal day (P)20 and P90. Serial coronal sections were obtained through the brainstem. Half of the sections were immunolabeled with antiserum to tyrosine hydroxylase (TH). Cell counts were obtained in areas A8 (retrorubral nucleus, RRN), A9 (substantia nigra, SN), and A10 (ventral tegmental area, VTA). The counts were analyzed with repeated measures analysis of variance followed by individual comparisons among group means. In A8, weaver homozygotes did not differ significantly from wild-type controls at P20, whereas there was a significant difference of 56% at P90. In A9, weaver homozygotes differed significantly from wild-type mice by 42% at P20 and by 69% at P90. The decrease in cell number between P20 and P90 in weaver homozygotes was 54%. In A10, weaver homozygotes did not differ significantly from wild-type controls at P20, whereas there was a significant difference of 26% at P90. Cell numbers in all three areas of heterozygotes did not differ significantly from wild-type control values at either age point. These findings demonstrate that by three months of age homozygous weaver mutants exhibit nerve cell losses in all three areas of the mesencephalic DA cell system. Such losses account for the DA deficiency seen in striatal, limbic and cortical projection fields.

Afterhyperpolarization (AHP) following single or short trains of spikes in rubrospinal neurones (RN neurones) of the cat has been studied with intracellular recording techniques. The AHP amplitude was potential dependent; it increased with depolarization and decreased with hyperpolarization and had an extrapolated reversal potential about 20 mV below resting membrane potential. The AHP was associated with an increase in the membrane conductance and it was concluded that the AHP is primarily caused by an increase in membrane conductance to potassium ions. The time course of the conductance change underlying the AHP was measured with short current pulses and calculated from the AHP voltage. The AHP following a single spike was conditioned at different interspike intervals by a preceding spike (or several spikes). In many RN neurones the AHP (conductance) following a spike added approximately linear to that generated by a preceding spike. In most cells, however, the AHP following a spike was instead depressed by a preceding spike. The summation of AHPs increased progressively, while the depression appeared to be already maximal with one preceding spike. The depression was then approximately constant for interspike intervals less than the AHP duration. It will be shown in a following paper that these properties of the AHP are reflected in the behaviour of the repetitive discharge evoked by constant current pulses in the same neurones.

Acetylcholine (ACh) is suggested to be a neurotransmitter in cerebellothalamic and cerebellorubral fibres. We therefore measured choline acetyltransferase (CAT) activity in the cat red nucleus after lesion of the contralateral cerebellum. Decreased CAT activity was obtained (-37%), specifically localized in the red nucleus and particularly in its rostral parvocellular part. Since the red nucleus of the cat displays rather low CAT levels, the possible cholinergic input seems not be sufficient to explain the powerful action of the cerebellum on red nucleus magnocellular neurons. As glutamic acid (Glu) may be the neurotransmitter of many excitatory systems in the brain, we measured high affinity Glu uptake in the red nucleus after lesions of the contralateral cerebellum. Results showed a decreased (-35%) Glu activity restricted to the red nucleus and particularly in its caudal magnocellular part.

Separate series of adult rats were subjected to unilateral high cervical and low thoracic section of the rubrospinal tract and sacrificed 1-30 (cervical series) and 3-100 days (thoracic series) later. Local cerebral glucose utilization ([14C]2-DG method of Sokoloff et al.) was determined in the red nucleus and in the inferior colliculus, nucleus interpositus and sensorimotor cortex of both sides in operates and controls. Although severe atrophy of rubral neurons follows cervical tractotomy while reversible chromatolytic alterations occur after thoracic lesions, glucose utilization did not differ in the red nucleus of operated and control rats. However, glucose utilization increased slightly in the inferior colliculus of all operated animals, a finding of indeterminate significance. The failure of axotomized intrinsic neurons of red nucleus and their surround to show altered glucose utilization stands in sharp contrast to the marked increase which occurs in cranial nerve nuclei after axotomy of their contained extrinsic neurons. The data are held to constitute another indication that there is a fundamental difference in the metabolic responses of extrinsic and intrinsic mammalian neurons to axotomy and may support the contention that, in mammals, the axon reaction of intrinsic neurons is fundamentally different from that of extrinsic nerve cells. This difference may have significance for failure of axon regeneration in mammalian CNS.(ABSTRACT TRUNCATED AT 250 WORDS)

3-acetylpyridine (3-AP) given in one single dose near to LD50 causes hydropic-vacuolic degeneration and cellular necrosis of varying degree and extension in the red nucleus of white rats in more than 50 percent of the cases. Less frequently similar alterations have been observed in some neighbouring nuclei of the mesencephalon. The regressive damage is remarkably rare if a smaller dose of 3-AP is given and none if the animal is pretreated with nicotine amide. Ultrastructurally the damage of mitochondria is dominant, they might be transformed to cystic lamellar figures. One reason of the enhanced vulnerability to antimetabolite is the outstanding enzyme activity of the red nucleus. In some of the intoxicated animals the six oxidative enzymes tested histochemically show different type of alterations which probably are related to the regressive damage.

Natural sleep entails an increase of the RNA concentration and absolute content (per 1 cell) in the gliocytes of the locus coeruleus and a decrease of the protein absolute content in the neuron cytoplasm. Within 24-hr REM-sleep deprivation, the protein and RNA concentrations remained constant in neurons and gliocytes whereas the RNA absolute content decreased in neurons; the 48-hr REM-sleep deprivation increased the protein concentration in neurons and glia (exceeding the wakefulness control values); protein absolute content in gliocytes increased as well, RNA concentration and absolute content remained equal to the control in neurons and in glia. In these respects the locus coeruleus differs from the supraoptic and red nucleus studied previously under the same conditions.

During hypothermia induced in ground squirrels by the halothane-heliox method, 2-deoxyglucose uptake of a white matter structure, the brachium conjunctivum, increased relative to the surrounding gray matter structures. The possibility of 2-deoxyglucose uptake by glial as well as neuronal elements in the brachium conjunctivum and the implications of this observation for the use of optical density ratios is discussed.

Following unilateral kainic acid lesioning of neuronal cell bodies in the cat red nucleus (RN), a large decrease in glutamic acid decarboxylase (GAD) activity was detected in the injected RN, compared to the RN from control, non-injected animals. Using GAD immunohistochemistry, reactive perikarya were visualized dorsolaterally to the rostral part of the nucleus as well as within the RN proper. Taken together, these results point to the existence of an intrinsic GABAergic innervation in the RN area of the cat. The GABAergic neurons characterized here might thus correspond to the inhibitory interneurons previously detected electrophysiologically as a putative source of GABA for large-sized neurons of the RN.

Fourteen kittens, 7--10 weeks of age, were injected with [3H]leucine 0.5--24 h before sacrifice 1--30 days after unilateral high cervical rubrospinal tractotomy. Histoautoradiographs of the red nuclei were prepared and counterstained with thionin. Axon reaction, evident histologically 24 h after surgery, was manifested by central chromatolysis or diffuse cytoplasmic chromophobia. Partial reversion toward a normal cytologic appearance was apparent 10--30 days postoperatively. Nucleolar and nuclear shrinkage and cytoplasmic atrophy were conspicuous accompaniments of axon reaction in rubral neurons. Expressed per cell the radioactivity of axotomized rubral nerve cells was consistently less than controls in animals surviving operation from 5 to 30 days. The data indicate that axon reaction in red nucleus is regressive in character and early associated with diminished protein synthesis. The frequently regressive nature of axon reaction in intrinsic neurons, such as those of red nucleus, probably is important in accounting for failure of regeneration of many mammalian CNS fiber tracts after injury.

Either the sialic acid precursor N-[3H]acetyl-D-mannosamine ([3H]ManNAc) or horseradish peroxidase (HRP) or both were injected into the mesencephalon of 15 rats. Localisation of radioactivity was performed by light microscopic autoradiography and HRP was histochemically localised with 3,3'-diaminobenzidine. After injection of either [3H]ManNAc or HRP, cell bodies in the central cerebellar nuclei were filled with radioactivity or HRP respectively. The distribution over contra- and ipsilateral interposed, dentate and fastigial nuclei of [3H]ManNAc-labelled and HRP-labelled cells is similar for both types of labelling. Since HRP is known to be transported retrogradely, a similar localisation of radioactivity-accumulating cells and HRP-filled cell bodies in the central cerebellar nuclei, lead us to the conclusion that radioactively-labelled ManNAc or its derivatives are also retrogradely transported in this system. There seem to be two possibilities for label uptake in afferents axons: (1) the uptake into axons which are disrupted by the injection needle and (2) the uptake by intact nerve endings or axons. After injection of [3H]ManNAc, the neuropil of central cerebellar nuclei shows a higher concentration of silver grains than the labelled cell bodies. This could be due to labelled dendrites caused by retrograde transport.

In an attempt to identify putative neurotransmitters of rubrospinal neurons, immunocytochemical procedures were utilized in combination with retrograde tracing techniques in 15 adult male rats. Following injections of horseradish peroxidase (HRP) or wheat germ agglutinin conjugated to HRP (WGA-HRP) into the spinal cord, midbrain sections were processed with a combined procedure that allowed visualization of both the retrograde tracer and one or more antigens including glutamate, glutaminase, and glutamatic acid decarboxylase (GAD). Initial colocalization studies demonstrated that glutamatelike and glutaminaselike immunoreactivities were cocontained within the same neurons. Following injections of HRP or WGA-HRP into the spinal cord approximately 53% of retrogradely labeled neurons contained glutamate immunoreactivity. Triple-labeling experiments indicated that glutamatelike immunoreactivity was colocalized with glutaminase immunoreactivity in retrogradely labeled rubrospinal neurons. Retrogradely labeled neurons did not contain GAD immunoreactivity. Moreover, triple labeling experiments verified that glutamatelike immunoreactive retrogradely labeled cells did not cocontain GAD immunoreactivity. These studies demonstrate that glutamate and its synthesizing enzyme, glutaminase, are present in some rubrospinal neurons and raise the possibility that a component of the rubrospinal projection may be glutamatergic. GAD, on the other hand, is not present in rubrospinal neurons. This finding supports the hypothesis that GABAergic neurons play a role as interneurons in the red nucleus.

L-DOPA (L-3,4-dihydroxyphenylalanine) immunoreactivity was demonstrated in neurons of the cat ventral midbrain and its projection areas, using an immunohistochemical method in conjunction with a newly developed highly specific anti-L-DOPA serum. L-DOPA-immunoreactive (IR) neurons were found in the substantia nigra, retrorubral area and ventral tegmental area of Tsai. L-DOPA-labeled fibers and terminals were hardly detectable in the nigrostriatal pathway and in the caudate nucleus which showed very intense dopamine-immunoreactivity. In contrast, many short labeled processes were detectable in the central amygdala and, although very few in number, in the entorhinal cortex.