Effects of KCl-induced depolarization on the GABA concentration in the corpus striatum and in the substantia nigra

Summary The effects of potassium ion depolarization on the concentration, the synthesis and the utilization of GABA as well as the effects on the concentrations of glutamine, dopamine (DA) and DOPAC in the substantia nigra and in the corpus striatum of the rat were investigated.An intranigral KCl injection did not influence the GABA concentration or the synthesis and utilization of GABA in the substantia nigra. Following an intrastriatal KCl injection the GABA concentration increased and the glutamine concentration decreased in the corpus striatum. A marked increase in the DA concentration in the corpus striatum was seen following an intranigral KCl injection, whereas the DA concentration decreased and the DOPAC concentration increased following an intrastriatal KCl injection. It is concluded that the GABA in the substantia nigra and in the corpus striatum is affected differently by potassium ions, perhaps due to the different cellular localization of GABA in the two structures. The increase of GABA in the corpus striatum was accompanied by a decreased glutamine concentration indicating that glutamine might be a precursor of GABA.     

Effects of muscimol and picrotoxin on single unit activity of substantia nigra neurons

Intravenous administration of the GABA agonist, muscimol, caused dose-dependent increases in the unit activity of substantia nigra pars compacta (dopamine) neurons and an inhibition of nigral pars reticulata cells. The depressant effects of the drug upon reticulata neurons were reversible by subsequent administration of the GABA antagonists, picrotoxin and bicuculline HCl. However, the stimulatory effects of i.v. muscimol upon dopamine neurons were not abolished by these agents. Intravenous administration of picrotoxin alone caused only moderate increases in the activity of dopamine neurons (31% over baseline at 7.0 mg/kg), but markedly stimulated the firing of pars reticulata cells (154% over baseline at 7.0 mg/kg). In spite of the stimulation of dopamine neurons after i.v. muscimol, microiontophoresis of GABA and muscimol could inhibit the firing of both pars compacta and pars reticulata cells, although the reticulata neurons were much more sensitive to the inhibitory actions of these agents than the dopamine neurons. Considered together, these studies suggest that a population of neurons in the substantia nigra pars reticulata have the capacity to be more affected by a major GABA input to the nigra than the pars compacta dopamine neurons. The results further suggest that if the dopamine cells are regulated by GABAergic neurons of the striatonigral pathway, their regulation must be indirect and could involve a second inhibitory neuron within the nigra.     

Pedunculopontine-evoked excitation of substantia nigra neurons in the rat

The effects of electrical stimulation of the nucleus tegmenti pedunculopontinus on the unitary activity of identified neurons of the rat substantia nigra were studied. The experiments were carried out in intact rats as well as in animals bearing either chronic bilateral electrolytic lesions of the deep cerebellar nuclei or an acute lesion of the ipsilateral subthalamic nucleus. Excitation of both compacta and reticulata cells of the substantia nigra (many of the latter being output neurons since they are antidromically activated from the superior colliculus) was the predominant response recorded. Two types of excitations could be distinguished. The first was a direct orthodromic excitation (latency 2.9 ± 1.6ms; duration3.7 ± 1.9ms). The second was a sparse and less pronounced activation (latency 5.2 ± 1.8ms; duration13.0 ± 3.0ms). These two types of excitation were the only responses recorded in intact rats (10/51, 19.6%, orthodromic and 10/51, 19.6%, diffuse activation). When the cerebellar nuclei were destroyed 7–21 days prior to the recording, both excitations were still found (10/59, 16.9% and 15/59, 25.4%, respectively), whereas a minority (3/59, 5.0%) of neurons were inhibited. Conversely, when the subthalamic nucleus was lesioned the orthodromic response was still present (9/42, 21.4%) whereas the occurrence of the diffuse excitation greatly decreased (3/42, 7.1%) and a greater number of inhibitions (6/42, 14.2%) appeared. A small population of cells (12/85, 14.1%) were excited from the contralateral pedunculopontine nucleus either by the orthodromic or by the diffuse excitation. The total number of nigral neurons antidromically activated from the ipsilateral pedunculopontine nucleus was 9/152 (5.9%). The results provide evidence that the nucleus tegmenti pedunculopontinus gives a dual excitatory input to the substantia nigra either through a probable direct connection or through a polysynaptic pathway via the subthalamic nucleus. A few cells from both parts of the substantia nigra, in turn, project back to the nucleus tegmenti pedunculopontinus. In addition, our data give further support to the view that output fibers from the deep cerebellar nuclei do not synapse in the substantia nigra in the rat.     

The effect of long-term amphetamine administration on the activity of dopaminergic neurons of the substantia nigra

Spontaneously active neurons from the pars compacta region of the substantia nigra were studied in chloral hydrate anesthetized, immobilized rats which had been pretreated with daily intraperitoneal saline or d-amphetamine injections according to several schedules. The spontaneous firing rates of these neurons were not altered by any of the pretreatment regimens. In addition, the intravenous dose of d-amphetamine sulfate which was required to reduce the spontaneous firing rate of these cells by at least 50% (threshold dose) was not modified by amphetamine pretreatment. There was a significant positive linear correlation between the threshold dose and the initial spontaneous firing rate of the neurons from animals pretreated with multiple saline injections, but not from those receiving long-term d-amphetamine. These results demonstrate that there are no alterations in the sensitivity of dopaminergic neurons to amphetamine under conditions known to produce the enhanced effects of the drug on motor behaviors which develop during multiple injections.     

Relationship between the presence of dopaminergic neurons and GABA receptors in substantia nigra: Effects of lesions

Submaximal destruction of nigrostriatal dopaminergic projections resulted in a significant (25%) decrease in specific GABA binding in substantia nigra; under these conditions, striatal tyrosine hydroxylase activity was 15–44% of control. In rats with lesions which caused maximal destruction of nigrostriatal dopamine neurons (striatal tyrosine hydroxylase was less than 15% of control), specific GABA binding in substantia nigra was apparently not different from that obtained in intact controls. Two distinct processes may be occurring in response to the destruction of dopamine neurons: (1) the loss of GABA binding sites physically associated with nigral dopamine neurons; and (2) an increase in nigral GABA receptors associated with non-dopaminergic neurons. The latter process may result from a decrease in nigral GABA transmission secondary to the complete loss of dopaminergic synaptic activity in striatum.     

Sex difference in the turnover of GABA in the rat substantia nigra

Summary The turnover of GABA (estimated from the post-mortem accumulation of GABA), and the activity of glutamic acid decarboxylase and GABA transaminase, along with the saturation of both enzymes by cofactor pyridoxal phosphate, were studied in the substantia nigra of rats of both sexes. Although no sex differences were found in the in vitro measured characteristics of both enzymes involved in GABA metabolism, the turnover of GABA was greater in males. This finding is consistent with our previous reports showing the greater resistance of male rats to GABA-related convulsions.     

Hyperpolarizing action of baclofen on neurons in the rat substantia nigra slice

Rabbit carotid bodies were excised and the membrane potentials (MPs) of the glomus cells were recorded in vitro. Upstream injections of dopamine (DA; 25-250 micrograms) elicited a slow, long lasting depolarization in 76% of the cells studied. The remaining 24% were hyperpolarized by DA. Regardless of polarity, the DA-induced potential was associated with a significant increase in input resistance (Ro). On the other hand, changes in MP which were produced artificially by passing direct current through the recording electrode had little effect on Ro. It is concluded that DA alters the MP and Ro of glomus cells and that this may serve to modulate synaptic excitability within the carotid body.     

Low doses of ethanol inhibit the firing of neurons in the substantia nigra, pars reticulata: a GABAergic effect?

The intravenous administration of relatively low doses of ethanol (0.25–2.00 g/kg) produced a dose-dependent inhibition of the firing rate of the neurons located in the substantia nigra, pars reticulata (PR neurons). This effect was eliminated both by picrotoxin and bicuculline, two blockers of γ-aminobutyric acid (GABA) transmission, and potentiated by muscimol (a direct GABA agonist) and diazepam (a representative of the benzodiazepine class which facilitate GABA transmission). The specific benzodiazepine antagonist, Ro 15-1788, blocked the potentiating effect of diazepam on the ethanol effect but failed to antagonize ethanol-induced inhibition of the firing rate of the neurons. These results indicate that ethanol might inhibit the firing of PR neurons through a GABAergic mechanism. Moreover, since PR neurons are thought to exert an inhibitory control on nigral dopaminergic neurons, it is suggested that the depression of the activity of such inhibitory interneurons may be responsible for ethanol-induced stimulation of dopaminergic activity.     

Evidence for impaired GABAergic activity in the substantia nigra of amygdaloid kindled rats

The activity of the GABA-synthesizing enzyme glutamate decarboxylase (GAD) was determined in synaptosomal fractions from 12 brain regions of amygdaloid kindled rats. The only significant difference in regional GAD activities between kindled animals and sham-operated controls was a 40% decrease of GAD activity in the substantia nigra. The data suggest that impaired GABAergic function in the nigra may be involved in the initiation and propagation of amygdaloid-kindled seizures.     

Removal of the telencephalon spares turning induced by injection of GABA agonists and antagonists into the substantia nigra

Unilateral injections into the substantia nigra of GABA agonists or the GABA-related agents muscimol (5–50 ng), baclofen (10–50 ng) or β-aminobutyric acid (25–50 μg) induced intensive turning in the direction contralateral to the injected hemisphere. Intranigral injection of the GABA antagonist picrotoxin (100–500 ng) led to ipsiversive turning. Surgical removal of the whole telencephalon (including the neocortex, hippocampus, striatum, septal nuclei and amygdalae) influenced neither direction nor magnitude of the turning responses induced by these various drugs. A GABAergic system in the substantia nigra can, therefore, cause turning independent of telencephalic structures.     

Evidence for GABA receptors on serotonergic afferent terminals in the substantia nigra and on nigral efferent projections to the caudal mesencephalon

Changes in [3H]GABA binding in rat substantia nigra (SN) were examined after (1) hemitransections at various levels posterior to SN and (2) 5, 7-dihydroxytryptamine (5, 7-DHT) induced lesions of the dorsal or median raphe or SN. Hemitransections 2 mm caudal to SN resulted in a 52% decrease in specific high affinity [3H] GABA binding in SN when measured 2-3 weeks postoperatively. No significant change in nigral GABA binding was found in rats with hemitransections placed at a postcollicular level(through the rostral pons). 5, 7-DHT placements in median raphe, or directly into SN, caused a 27% decrease in specific [3H] GABA binding in SN. 5, 7-DHT lesions of the dorsal raphe did not significantly affect the binding of [3H]GABA in SN. These data suggest that a significant protion of GABA receptors in SN may be located presynaptically on axon terminals of serotonergic projections from the median raphe to SN. Other pathways between the SN and the caudal mesencephalon also appear to contain GABA receptors; these are likely to be efferent nigral projections to the mesencephalic reticular formation.     

Effects of dopaminergic cell degeneration on electrophysiological characteristics and GAD65/GAD67 expression in the substantia nigra: different action on GABA cell subpopulations.

The motor disturbances occurring in Parkinson's disease have been partially attributed to a hyperactivity of gamma-aminobutyric acid (GABA)-ergic nigral cells largely in the substantia nigra pars reticulata (SNr) secondary to the degeneration of dopaminergic nigrostriatal neurons. However, some aspects of this response remain unclear. In this work, different electrophysiological and neurochemical parameters were studied in GABAergic cells of the SN after unilateral nigrostriatal dopaminergic lesion using 6-hydroxydopamine injection in rats. Our data showed that 1) the SN under normal conditions contains different subsets of GABAergic cells according to their firing pattern and glutamic acid decarboxylase mRNA levels, and 2) the response of these GABAergic cell subgroups was different after the ipsi- and contralateral dopaminergic cell degeneration. These findings indicate a complex regulation of nigral GABAergic activity after nigrostriatal dopaminergic degeneration that probably involves local mechanisms, the nigro-striato-nigral loop, as well as interhemispheric mechanisms whose anatomical basis remains unstudied.     

Seizure suppression in kindling epilepsy by grafts of fetal GABAergic neurons in rat substantia nigra

Compared with studies on models of neurodegenerative diseases, considerably less work has been performed with neural grafts in experimental epilepsy. The potential value of this approach, however, is already shown by evidence that noradrenergic grafts implanted bilaterally into the hippocampus or amygdala-piriform cortex can suppress seizure development in the kindling model of temporal lobe epilepsy. We previously showed that amygdala kindling results in a significant decrease of GABA and its synthesizing enzyme glutamate decarboxylase in substantia nigra (SN), i.e., a region thought to be critically involved in seizure propagation in various models of epilepsy. Thus, transplantation of fetal GABAergic neurons into SN might be an effective means of permanently blocking seizure generalization in kindling epilepsy and probably also other types of epilepsy. To test this hypothesis, three groups of female Wistar rats (n = 10 per group) were kindled by electrical stimulation via a bipolar electrode in the basolateral amygdala. After all rats were fully kindled, one group was implanted with GABA-rich cells prepared from the striatal eminence of Wistar rat fetuses at embryonic day 14. The striatal neurons were bilaterally microinjected at various sites over the anterior-posterior axis of the SN, aimed at the pars reticulata. The second group received microinjections of spinal cord cell preparations, whereas the third group received microinjections of cell-free medium only. In all rats, the threshold for focal discharges (afterdischarge threshold [ADT]) as well as afterdischarge duration and severity and duration of seizures occurring at ADT current were determined once weekly before and after transplantation. Eleven to 12 weeks following transplantation, the rats were killed, and location and integration of grafts were examined by immunohistological methods. Rats with GABAergic grafts in SN exhibited a significant increase in ADT and marked reduction in seizure severity compared with pretransplantation values, whereas no such alteration was seen in the other groups. However, the seizure-suppressing effect of GABAergic grafts was not permanent but slowly disappeared over the weeks after transplantation. Although the data indicate that intranigral transplantation of GABA-producing cells is no effective means of inducing long-lasting anticonvulsant effects in experimental epilepsy, this approach may be an initial step to develop more efficient strategies for seizure suppression. J. Neurosci. Res. 51:196–209, 1998. © 1998 Wiley-Liss, Inc.     

Impact of expected value on neural activity in rat substantia nigra pars reticulata

The substantia nigra pars reticulata (SNr) is thought to serve as the output of the basal ganglia, whereby associative information from striatum influences behavior via disinhibition of downstream motor areas to motivate behavior. Unfortunately, few studies have examined activity in SNr in rats making decisions based on the value of predicted reward similar to those conducted in primates. To fill this void, we recorded from single neurons in SNr while rats performed a choice task in which different odor cues indicated what reward was available on the left or on the right. The value of reward associated with a leftward or rightward movement was manipulated by varying the size of and delay to reward in separate blocks of trials. Rats were faster or slower depending on whether the expected reward value was high or low, respectively. The number of neurons that increased firing during performance of the task outnumbered those that decreased firing. Both increases and decreases were modulated by expected value and response direction. Neurons that fired more or less strongly for larger reward tended to fire, respectively, more or less strongly for immediate reward, reflecting their common motivational output. Finally, value selectivity was present prior to presentation of cues indicating the nature of the upcoming behavioral response for both increasing- and decreasing-type neurons, reflecting the internal bias or preparatory set of the rat. These results emphasize the importance of increasing-type neurons on behavioral output when animals are making decisions based on predicted reward value.     

Influence of electrical stimulation of the substantia nigra on spontaneous activity of raphe neurons in the anesthetized rat

Recent studies of afferent connections of the anterior raphe using the horseradish peroxidase technique have demonstrated a major projection originating in the substantia nigra (SN). The present acute electrophysiological study examined the influence of stimulation of this afferent on the activity of individual neurons in the raphe of the posterior midbrain and anterior pons (n=51), and of a control group of cells (n=15) located 2 mm lateral to the raphe. The predominant effect of SN stimulation at 0.1–1.0 mA, 1 Hz or 10 Hz, was suppression of raphe activity, with 63% of the cells showing cessation of firing following SN pulses and only 8% showing excitation. The average duration of suppression was 200 msec at 1 Hz and 38 msec at 10 Hz. In contrast, 40% of the lateral cells were excited, with 27% of the cells showing suppression. The mean duration of total suppression of lateral cell firing was 61 and 17 msec at 1 and 10 Hz, respectively. The results from the raphe cells are consistent with recent reports of stimulation of other forebrain and brainstem afferents to the raphe in which suppression of raphe activity was the main effect.     

Distinct commissural pathways are involved in the enhanced release of dopamine induced in the contralateral caudate nucleus and substantia nigra by unilateral application of GABA in the cat thalamic motor nuclei

The effects of diencephalic or telencephalic commissural sectioning on the changes in [3H]dopamine ([3H]DA) release from nerve terminals (in the caudate nucleus, CN) and dendrites (in the substantia nigra, SN) of the two nigro-striatal dopaminergic pathways induced by the application of GABA (10(-5) M, 30 min) into the left ventralis medialis (VM) or ventralis lateralis (VL) thalamic nuclei were investigated. Experiments were performed in halothane-anesthetized cats implanted with push-pull cannulae in both CN and SN. In unlesioned cats, GABA application into the left VM-VL increased [3H]DA release in both CN and in the contralateral SN confirming previous results. Sectioning of the thalamic massa intermedia only blocked the GABA-induced increase in [3H]DA release in the contralateral SN, the responses in both CN being preserved. Sectioning of the rostral part of the corpus callosum only prevented the GABA-induced increase in [3H]DA release in the contralateral CN, whereas [3H]DA release in the ipsilateral CN and in the contralateral SN was still enhanced. These results suggest that changes in [3H]DA release evoked in both CN and in the contralateral SN by GABA application into the left VM-VL might involve different mechanisms: those observed in the CN result from potent pre-synaptic influences mediated by the bilateral cortico-striatal projections; those induced in the contralateral SN are due to other types of messages involving or passing through the thalamic massa intermedia.     

GIRK2 expression in dopamine neurons of the substantia nigra and ventral tegmental area

G-protein-regulated inward-rectifier potassium channel 2 (GIRK2) is reported to be expressed only within certain dopamine neurons of the substantia nigra (SN), although very limited data are available in humans. We examined the localization of GIRK2 in the SN and adjacent ventral tegmental area (VTA) of humans and mice by using either neuromelanin pigment or immunolabeling with tyrosine hydroxylase (TH) or calbindin. GIRK2 immunoreactivity was found in nearly every human pigmented neuron or mouse TH-immunoreactive neuron in both the SN and VTA, although considerable variability in the intensity of GIRK2 staining was observed. The relative intensity of GIRK2 immunoreactivity in TH-immunoreactive neurons was determined; in both species nearly all SN TH-immunoreactive neurons had strong GIRK2 immunoreactivity compared with only 50-60% of VTA neurons. Most paranigral VTA neurons also contained calbindin immunoreactivity, and approximately 25% of these and nearby VTA neurons also had strong GIRK2 immunoreactivity. These data show that high amounts of GIRK2 protein are found in most SN neurons as well as in a proportion of nearby VTA neurons. The single previous human study may have been compromised by the fixation method used and the postmortem delay of their controls, whereas other studies suggesting that GIRK2 is located only in limited neuronal groups within the SN have erroneously included VTA regions as part of the SN. In particular, the dorsal layer of dopamine neurons directly underneath the red nucleus is considered a VTA region in humans but is commonly considered the dorsal tier of the SN in laboratory species.     

Inhibition of neuronal activity of the substantia nigra by noxious stimuli and its modification by the caudate nucleus

In urethane-anesthetized rats discharges of neurons of substantia nigra, pars compacta (SNC) were recorded extracellularly after natural somatic sensory stimulation and electrical stimulation of peripheral sensory nerves.

(1) Among different modalities of somatic sensory stimulation tested, noxious stimuli were effective in reducing spontaneous discharges of SNC neurons. The inhibition appeared with a concomitant increase of spike amplitude. The same inhibitory effect was obtained by stimulating the sciatic nerve (SC) repetitively. In response to single shock stimulation of the SC the inhibition occurred at an average latency of 39.6 msec (S.E. 1.6 msec) and lasted for 221.6 msec on average (S.E. 10.8 msec).

(2) The SC-induced inhibition of SNC neurons failed to reliably block orthoand antidromic discharges evoked from the caudate nucleus (Cd).

(3) In rats with the Cd lesioned the SC-induced inhibition was longer lasting than in controls. When the Cd was stimulated concurrently with SC stimulation, the inhibition from the SC was weakened.

(4) In a majority of SNC neurons, their inhibition by SC stimulation was antagonized by intravenous injection of haloperidol.