The novel delta opioid receptor agonist UFP-512 dually modulates motor activity in hemiparkinsonian rats via control of the nigro-thalamic pathway

The present study aimed to characterize the ability of the novel delta opioid peptide (DOP) receptor agonist H-Dmt-Tic-NH-CH(CH₂–COOH)-Bid (UFP-512) to attenuate motor deficits in 6-hydroxydopamine (6-OHDA) hemilesioned rats. Lower doses (0.1–10 μg/kg) of UFP-512 administered systemically (i.p.) stimulated stepping activity in the drag test and overall gait abilities in the rotarod test whereas higher doses (100–1000 μg/kg) were ineffective or even worsened Parkinsonism. Microdialysis coupled to an akinesia test (bar test) was then used to determine the circuitry involved in the motor actions of UFP-512. An antiakinetic dose of UFP-512 (10 μg/kg) decreased GABA in globus pallidus (GP) as well as GABA and glutamate (GLU) release in substantia nigra reticulata (SNr). On the other hand, a pro-akinetic dose (1000 μg/kg) of UFP-512 increased pallidal GABA, simultaneously producing a decrease in GABA and an increase in nigral GLU release. Moreover, to test the hypothesis that changes in motor behavior were associated with changes in nigro–thalamic transmission, amino acid release in ventromedial thalamus (VMTh, a target of nigro–thalamic GABAergic projections) was also measured. The anti-akinetic dose of UFP-512 reduced GABA and increased thalamic GLU release while the pro-akinetic dose increased GABA without affecting thalamic GLU release. Finally, regional microinjections were performed to investigate the brain areas involved in motor actions of UFP-512. UFP-512 microinjections into GP increased akinesia whereas UFP-512 microinjections into SNr reduced akinesia. Furthermore, the selective DOP receptor antagonist naltrindole (NTD) increased akinesia when injected into either area, GP being more sensitive. We conclude that UFP-512, depending on dose, improves or worsens motor activity in hemiparkinsonian rats by acting differentially as a DOP receptor agonist in SNr and a DOP receptor antagonist in GP, ultimately decreasing or increasing the activity of nigro–thalamic GABAergc output neurons, respectively.     

Synaptically released GABA activates both pre- and postsynaptic GABA(B) receptors in the rat globus pallidus

The globus pallidus (GP) contains abundant GABAergic synapses and GABA(B) receptors. To investigate whether synaptically released GABA can activate pre- and postsynaptic GABA(B) receptors in the GP, physiological recordings were performed using rat brain slice preparations. Cell-attached recordings from GABA(A) antagonist-treated preparations revealed that repetitive local stimulation induced a GABA(B) antagonist-sensitive pause in spontaneous firings of GP neurons. Whole cell recordings revealed that the repetitive stimulation evoked fast excitatory postsynaptic potentials followed by a slow inhibitory postsynaptic potential (IPSP) in GP neurons. The slow IPSP was insensitive to a GABA(A) receptor antagonist, increased in amplitude with the application of ionotropic glutamate receptor antagonists, and was suppressed by the GABA(B) antagonist CGP55845. The reversal potential of the slow IPSP was close to the potassium equilibrium potential. These results suggest that synaptically released GABA activated postsynaptic GABA(B) receptors and induced the pause and the slow IPSP. On the other hand, in the neurons that were treated to block postsynaptic GABA(B) responses, CGP55845 increased the amplitudes of repetitive local stimulation-induced GABA(A)-mediated inhibitory postsynaptic currents (IPSCs) but not the ionotropic glutamate-mediated excitatory postsynaptic currents. Moreover, the GABA(B) receptor specific agonist baclofen reduced the frequency of miniature IPSCs without altering their amplitude distributions. These results suggest that synaptically released GABA also activated presynaptic GABA(B) autoreceptors, resulting in decreased GABA release in the GP. Together, we infer that both pre- and postsynaptic GABA(B) receptors may play crucial roles in the control of GP neuronal activity.     

Stimulation of GABA release from the rat neostriatum and globus pallidus in vivo by corticotropin-releasing factor

This study conducted in vivo examined the changes in gamma-aminobutyric acid (GABA) release in push-pull perfusates of the caudate nucleus (CN) and the globus pallidus (GP) in response to corticotropin releasing factor (CRF). In the CN, CRF (10(-12), 10(-10), 10(-8) M) stimulated GABA release in a dose-related manner, the highest dose (10(-8) M) also potentiating the 25 mM K+-evoked response. The release of GABA in response to CRF (10(-8) M) was completely blocked by alpha-helical CRF9-41 (10(-6) M) which also attenuated the K+-evoked response to control K+-stimulated levels. In the GP, only the highest dose of CRF (10(-8) M) significantly stimulated GABA release, this dose also potentiating the K+-evoked release. Both responses were attenuated by the CRF receptor antagonist (10(-6) M). These results thus demonstrate that CRF can exert potent effects on GABA release within the rat neostriatum-pallidum by increasing the membrane excitability of GABA neurons/terminals and that such effects are mediated via receptors present on both the cell bodies/terminals of GABA-containing neurones. These effects of CRF suggest that the peptide may be an integral component of the neurochemical circuitry in the basal ganglia with relevance to the regulation of motor behaviour.     

Tissue levels and in vivo release of tachykinins and GABA in striatum and substantia nigra of rat brain after unilateral striatal dopamine denervation

Summary Brain tissue levels and in vivo release of substance P (SP) and neurokinin A (NKA) and GABA were measured bilaterally in striatum and substantia nigra of the rat, after a unilateral 6-hydroxydopamine lesion of the nigro-striatal dopamine pathway. Sham injected animals served as controls. The dopamine denervation decreased the tissue levels of SP in striatum (-38%) ipsilateral to the lesion and in substantia nigra both ipsi- (-54%) and contralateral (-38%) to the lesion. NKA was not significantly changed in the striatum, but decreased (like SP) in the substantia nigra both ipsi- (-50%) and contralateral (-40%) to the lesion. GABA tissue levels increased in the denervated striatum (+20%) and remained unchanged in substantia nigra at both sides. The extracellular levels of SP, NKA and GABA were measured with microdialysis in vivo at basal conditions and during stimulation with potassium administered locally via the microdialysis probe. The stimulated release of SP and NKA in the substantia nigra ipsilateral to the lesion was compared to in sham operated animals reduced with 39% and 64%, respectively, while no change in SP or NKA release was detected in the striatum. The basal release of GABA in the striatum was increased with 296% and with 76% during stimulation in the dopamine denervated striatum, while no change in GABA basal or stimulated release was detected in the substantia nigra. We suggest that the increased GABA release in the dopamine denervated striatum may be due to a decreased dopamine mediated inhibition of local GABA neurons. Furthermore, the decreased nigral release of SP and NKA ipsilateral to the lesion is suggested to be caused by an increased GABA inhibition in striatum of SP- and NKA-containing striato-nigral neurons.     

Role of globus pallidus and substantia nigra efferent pathways in striatally evoked head turning in the rat

Summary These studies have examined the role of brain areas that receive efferent projections from the globus pallidus (GP) and substantia nigra (SN) in producing the contralateral head turning evoked by unilateral electrical stimulation of the neostriatum in the conscious rat. Two parameters were studied: the latency for a 90 ° head turn and changes in the normal latency evoked by administration of the GABA drugs picrotoxin and muscimol in GP. Electrolesions in the ipsilateral ventromedial and centromedian thalamic nuclei had no effect on the head turn parameters. Although small electrolesions in the SN slowed, but did not abolish the head turn, it prevented the changes in the response latency brought about by GABA drugs in GP. Treatment with 6-hydroxydopamine, which partly destroyed the nigrostriatal dopamine neurones, had no effect on the head turn. Areas of the brainstem that receive basal ganglia efferents were lesioned. An electrolesion of the nucleus tegmenti pedunculopontinus had no effect on the head turn latencies. The head turning was abolished by a lesion in the lateral periaqueductal grey (PAG); a more rostral PAG lesion, on the same level as SN, was ineffective. It is concluded that the head turning is mediated by basal ganglia efferents which pass close to the nigra without synapsing and project to the PAG. The GABA-sensitive GP efferents which modulate the response, probably project to SN.     

In vivo release of [3H]gamma-aminobutyric acid in the rat neostriatum--I. Characterization and topographical heterogeneity of the effects of dopaminergic and cholinergic agents

The release of [3H]gamma-aminobutyric acid continuously synthesized from [3H]glutamine was studied in the striatum of halothane-anaesthetized rats superfused with a push-pull cannula. The levels of spontaneously released [3H]GABA were identical in all striatal regions examined, but were found to be higher at the junction between the striatum and the globus pallidus. Superfusion with a medium enriched in K+ ions induced a concentration-dependent increase in [3H]GABA release. Superfusion with a Ca2+-free medium did not affect the spontaneous outflow of [3H]GABA but sharply reduced the release of [3H]GABA evoked by 30 mM K+. Locally applied tetrodotoxin (50 microM) decreased slightly the spontaneous release of [3H]GABA (-22%). When acetylcholine (50 or 500 microM) was added to a superfusion medium containing eserine (50 microM), the spontaneous release of [3H]GABA was enhanced in the ventral but not in the dorsal region of the striatum. The local application of 2,3,4,5-tetrahydro, 7,8,-dihydroxy, 1-phenyl, 1-H, 3-benzazepine (10 microM), a dopaminergic agonist acting preferentially on D1 receptors increased the release of [3H]GABA in the dorsal striatum (+32%) but decreased it slightly (-19%) in the ventral striatum. 3-(2-(N-3 hydroxyphenylethyl)N-propylamino)ethyl-phenol (50 microM), a preferential D2 receptor agonist, decreased [3H]GABA release when it was applied dorsally (-23%) but not ventrally in the striatum. It is concluded that the regulation of the release of [3H]GABA by acetylcholine and dopaminergic drugs is different in the dorsal and ventral regions of the striatum. These differences may be related to the existence of subpopulations of GABA neurons and may well have functional implications as suggested by behavioural studies.     

Adenosine A(2A) receptor-mediated modulation of GABA and glutamate release in the output regions of the basal ganglia in a rodent model of Parkinson's disease

A target neuron of adenosine A(2A) receptor antagonists to exert anti-parkinsonian activities has been currently identified to be, at least in part, striatopallidal medium spiny neurons (MSNs). In the present study, we determine whether A(2A) receptor-mediated modulation is associated with changes in the release of GABA and glutamate in the substantia nigra pars reticulata (SNr), an output structure of the whole basal ganglia network, using in vivo microdialysis in a rat Parkinson's disease (PD) model. In 6-hydroxydopamine (OHDA)-lesioned rats compared with normal rats, basal extracellular GABA levels in the SNr show no change, whereas basal glutamate levels are significantly increased. Oral administration of the A(2A) receptor-selective antagonist (E-1,3-diethyl-8-(3,4-dimethoxystyryl)-7-methyl-3,7-dihydro-1-H-purine-2,6-dion (KW-6002) to 6-OHDA-lesioned rats at 1 mg/kg caused a marked and sustained increase of GABA and glutamate levels in the SNr. The increase of nigral glutamate by KW-6002 was abolished by a kainic acid-induced lesion of the globus pallidus (GP) or subthalamic nucleus (STN) in 6-OHDA-lesioned rats, whereas the increase of nigral GABA was completely blocked by the GP-lesion but only partially blocked by the STN-lesion. These results indicate that changes in neurotransmitter release in the SNr brought about by KW-6002 are largely attributable to blockade of A(2A) receptor-mediated modulation of striatopallidal MSNs. Thus, these actions of KW-6002 on striatopallidal MSNs may be the main mechanism for ameliorating PD by A(2A) antagonists.     

In vivo release of [3H]gamma-aminobutyric acid in the rat neostriatum--II. Opposing effects of D1 and D2 dopamine receptor stimulation in the dorsal caudate putamen

The effects of several dopaminergic agonists and antagonists on the spontaneous release of [3H]gamma-aminobutyric acid were investigated in the dorsal striatum of halothane-anaesthetized rats. A push-pull cannula was implanted and the tissue was superfused continuously with a physiological medium containing [3H]glutamine, the precursor of [3H]GABA. Drugs were added to the superfusion medium. 2-Amino,6,7-dihydroxy,1,2,3,4-tetrahydro-naphtalene (ADTN, a mixed D1 and D2 receptor agonist) and D-amphetamine (a drug that enhances the release of endogenous dopamine) increased the release of 3H-GABA. The effect of ADTN was blocked by a D1 antagonist [R-(+),8-chloro, 7-hydroxy,2,3,4,5-tetrahydro,3-methyl,5-phenyl,1-H,3-benzazepine (SCH 23390)] but not by a D2 antagonist (S-sulpiride). Furthermore the stimulation of D1 receptors either by 2,3,4,5-tetrahydro,7,8-dihydroxy,1-phenyl,1-H,3-benzazepine or by D-amphetamine in the presence of S-sulpiride also enhanced the release of [3H]GABA. On the other hand, a selective D2 receptor agonist (3-(2-(N-3-hydroxy-phenylethyl)N-propylamino)ethyl-phenol) decreased the release of [3H]GABA. This effect was blocked in the presence of S-sulpiride. By itself the D1 receptor antagonist (SCH 23390) decreased the release of [3H]GABA whereas the D2 receptor antagonist (S-sulpiride) had no effect. It was concluded that stimulation of D1 and D2 receptors produces opposing effects on the spontaneous release of [3H]GABA in the dorsal striatum. Stimulation of D1 receptors facilitates the release of [3H]GABA whilst stimulation of D2 receptors inhibits it. The effect of D1 receptor stimulation appears to be predominant, and endogenous dopamine may activate tonically the release of GABA through these receptors in our experimental conditions.(ABSTRACT TRUNCATED AT 250 WORDS)     

Opposing effects of dopamine D2 receptor stimulation on the spontaneous and the electrically evoked release of [3H]GABA on rat prefrontal cortex slices.

The spontaneous and the electrically evoked release of [3H]GABA were studied in vitro on slices of rat medial prefrontal cortex. The slices were preincubated with [3H]GABA and then superfused with a Krebs' solution. The superfusion with a Ca(2+)-free medium progressively increased the spontaneous [3H]GABA release and strongly decreased the electrically evoked release of [3H]GABA (-65%). The effects of three dopaminergic D2 receptor agonists (RU24926, lisuride and LY171555) were studied on both the spontaneous and the electrically evoked [3H]GABA release. The spontaneous release of [3H]GABA was increased by exposure to each of these three D2 agonists. RU24926 produced a dose-dependent increase from 10(-9) to 3 x 10(-8) M and the maximal effect was totally abolished by the dopaminergic D2 receptor antagonist sulpiride (10(-5) M). With lisuride a progressive increase of [3H]GABA release was observed and a plateau value was reached with concentrations between 10(-7) and 10(-6) M. These effects were totally reversed by 10(-5) M sulpiride. The dose-response relation for LY171555 was bell-shaped, with a maximal effect being obtained with 10(-9) M) LY171555. This effect decreased with a higher concentration (10(-8) M) and finally was no longer observed for 10(-7) M LY171555. The maximal increase induced by LY171555 was totally abolished by 10(-5) M sulpiride. In contrast, the electrically evoked release of [3H]GABA was inhibited by these three D2 agonists. The IC50 value of the inhibition was 4.1 x 10(-8) M for RU24926 and 2 x 10(-7) M for lisuride. Sulpiride (10(-5) M) totally abolished the effect of 10(-7) M RU24926. In the concentration range of lisuride examined, a 50% reduction of the lisuride inhibition was obtained in the presence of sulpiride (10(-5) M). The dose-response curve obtained with LY171555 had a U-shape, with a maximal inhibition reached with 10(-8) M, whereas no effect was observed with 10(-6) M. The inhibition induced by 10(-8) M LY171555 was completely antagonized by 10(-5) M sulpiride. The D2 agonist-induced inhibition of the electrically evoked release of [3H]GABA was mimicked by dopamine endogenously released by 10(-5) M amphetamine. This effect was reversed by 10(-5) M sulpiride. Our data provide further evidence for a dopaminergic control of GABA interneurons in the prefrontal cortex. This regulation implies the activation of D2 dopaminergic receptors. The possible mechanisms underlying the opposite effects of D2 agonists on the spontaneous and the electrically evoked release of [3H]GABA are discussed.     

Effects of somatostatin-14 on the in vitro release of [3H]GABA from slices of rat caudatoputamen

Slices (300 microns) of rat caudatoputamen were incubated in Krebs-Henseleit medium and loaded with [3H]glutamine, part of which was converted to [3H]GABA. This conversion takes place only in GABA-neurons most of which probably contribute to the striatonigral pathway. After a 24 min equilibration period, release of radioactivity was stimulated with veratridine (3.1-4 mumol/l) or K+ (15-25 mmol/l) in the absence or presence of somatostatin-14. From the radioactivity released [3H]GABA was separated by cationic exchange chromatography and measured. Somatostatin-14 affected the release of [3H]GABA in a manner which depended on its concentration as well as on the extent of stimulus-evoked release. Somatostatin-14 (1 nmol/l) enhanced the moderate release (2-4% of tissue content) elicited by veratridine (3.1 mumol/l) or K+ (20 mmol/l), but had no effect on the more pronounced release (5-8% of tissue content) elicited by veratridine (4 mumol/l) or K+ (25 mmol/l). Somatostatin-14 (10 nmol/l) had no effect on the moderate release of [3H]GABA, but diminished the pronounced one. Further experiments provided evidence that the somatostatin-14-induced enhancement was not brought about by a direct action on GABA-neurons but was probably indirect, i.e. mediated by other striatal neurons. In contrast, the diminution of the release of [3H]GABA caused by somatostatin-14 may be due to its direct action on releasing neurons. Two antisera against somatostatin lowered the pronounced release indicating that endogenous somatostatin may also enhance the release of [3H]GABA. In addition, endogenous somatostatin seems also to be able to diminish the release under certain experimental conditions.(ABSTRACT TRUNCATED AT 250 WORDS)     

GAT-3 transporters regulate inhibition in the neocortex

The role of GAT-3 transporters in regulating GABA(A) receptor-mediated inhibition was examined in the rat neocortex using an in vitro slice preparation. Pharmacologically isolated GABA(A) receptor-mediated responses were recorded from layer V neocortical pyramidal cells, and the effects of SNAP-5114, a GAT-3 GABA transporter-selective antagonist, were evaluated. Application of SNAP-5114 resulted in a reversible increase in the amplitude of an evoked GABA(A) response in most cells examined, although no effect on the decay time was observed. Examination of the spontaneous output of inhibitory interneurons revealed a reversible increase in the frequency and amplitude of spontaneous inhibitory synaptic currents as a consequence of GAT-3 inhibition. This effect of GAT-3 inhibition on spontaneous inhibitory events was action potential-dependent because no such increases were observed when SNAP-5114 was applied in the presence of TTX. These results demonstrate that GAT-3 transporters regulate inhibitory interneuron output in the neocortex. The increase in inhibitory interneuron excitability resulting from application of SNAP-5114 suggests that inhibition of GAT-3 transporter function results in a reduction in ambient GABA levels, possibly by a reduction in carrier-mediated GABA release via the GAT-3 transporter.     

Dynorphin exerts both postsynaptic and presynaptic effects in the Globus pallidus of the rat

The opioids contained in striato-pallidal axons are thought to play a significant role in motor control. We examined post- and presynaptic effects of the kappa (kappa)-receptor agonist dynorphin A (1-13) (DYN13) on the globus pallidus (GP) neurons in rat brain slice preparations using the whole cell recording method. DYN13 hyperpolarized and decreased the input resistance of approximately one-quarter of neurons examined. All of these DYN13-sensitive neurons had medium-sized somata, large aspiny dendrites and generated repetitive firing without strong accommodation. The hyperpolarization was blocked by barium and was independent of TTX and intracellular chloride levels. The hyperpolarization was also selectively blocked by the kappa-antagonist nor-binaltorphimine dihydrochloride but not by the mu- or delta-antagonists. These data suggested that DYN13 activates barium-sensitive potassium currents in some GP neurons. Low- and high-intensity stimulation of the neostriatum (Str) evoked long- and short-latency GABAergic responses, respectively. Previous data suggested that the long- and the short-latency responses were due to activation of the striato-pallidal axons and the local collaterals of pallido-striatal axons, respectively. DYN13 diminished the amplitude of both the short- and long-latency GABAergic responses in all the neurons tested. The effects of DYN13 on GABAergic postsynaptic responses were also selectively blocked by a kappa-antagonist. To investigate whether the effects were pre- or postsynaptic, the effects of DYN13 on spontaneous inhibitory postsynaptic potentials (IPSPs) and TTX-independent miniature-inhibitory postsynaptic currents (IPSCs) were examined. DYN13 decreased the frequency, but not the amplitude, of spontaneous IPSCs and calcium-dependent miniature-IPSCs. However, DYN13 did not alter the cadmium-insensitive miniature-IPSCs. These results suggested that DYN13 suppressed GABA release from presynaptic terminals. This possibility was tested using a paired-stimulation test. DYN13 reduced the probability of evoking IPSCs to the first stimulation and greatly increased the success probability to the second stimulus. The amplitude of successfully evoked IPSCs was not changed with DYN13. DYN13 did not affect the excitatory postsynaptic potentials (EPSPs) or the response to iontophoretically applied GABA and glutamate. Together, these results suggest that DYN released from striato-pallidal axons controls the activity of GP neurons 1) by directly hyperpolarizing a population of neurons and 2) by presynaptically inhibiting GABA release from striato-pallidal and intrapallidal terminals.     

The release of γ-aminobutyric acid during inhibition in the cat visual cortex

1. The release of gamma-aminobutyric acid (GABA) from the surface of the posterior lateral gyrus of the cerebral cortex was measured by a sensitive enzymic fluorimetric assay procedure. Experiments were performed with anaesthetized cats during resting conditions and during cortical inhibition produced by electrical stimulation of the brain surface or of the lateral geniculate nucleus (l.g.n.).2. The average resting release of endogenous GABA was 0.20 n-mole/ 7 min.cm(2) cortex; this was increased during stimulation of both the cortical surface (2.9 times resting release during monopolar stimulation and 7.4 times resting release during bipolar stimulation) and the l.g.n. (5.7 times resting release).3. Removal of calcium ions from the collection fluid did not affect the resting release of endogenous GABA but prevented the increase in GABA release normally evoked by stimulation of the cortical surface.4. The stimulus parameters used to increase the release of GABA also inhibited the glutamate-induced firing of single cells in the visual cortex and this inhibition was abolished in the absence of calcium ions.5. In three experiments the total amino acid content of cortical samples was examined using an amino acid analyser. With the exception of GABA, there were no significant differences between the rates of release of any other detected amino acids during periods with and without electrical stimulation of the cortex.6. It is suggested that since the release of GABA observed during inhibitory stimulation of the cortex is calcium-dependent and specific, it may originate from inhibitory nerve terminals in the cortex. The present findings support the view that GABA is a central inhibitory neurotransmitter.     

Calcium influx through presynaptic 5-HT3 receptors facilitates GABA release in the hippocampus: in vitro slice and synaptosome studies

Serotonin 5-hydroxytryptamine type 3 receptors (5HT3R) are Ca2+-permeant, non-selective cation channels that have been localized to presynaptic terminals and demonstrated to modulate neurotransmitter release. In the present study the effect of 5-HT on GABA release in the hippocampus was characterized using both electrophysiological and biochemical techniques. 5-HT elicited a burst-like, 6- to 10-fold increase in the frequency of GABAA receptor-mediated inhibitory postsynaptic currents (IPSCs) measured with whole-cell voltage-clamp recordings of CA1 neurons in hippocampal slices. When tetrodotoxin was used to block action potential propagation, the 5-HT-induced burst of IPSCs was still observed. Stimulation of hippocampal synaptosomes with 5-HT resulted in a significant increase in the amount of [3H]GABA released by hyperosmotic saline. In both preparations, the 5-HT effect was shown to be mediated by 5HT3Rs, as it was mimicked by the selective 5HT3R agonist m-chlorophenyl biguanide and blocked by the selective 5HT3R antagonist 3-tropanylindole-3-carboxylate hydrochloride. The 5HT3R-mediated increase in GABA release was blocked by 100 microM cadmium or by omitting Ca2+ in external solutions, indicating the Ca2+-dependence of the effect. The high voltage-activated Ca2+ channel blockers omega-conotoxin GVIA and omega-conotoxin MVIIC and 10 microM cadmium had no significant effect on the 5-HT3R-mediated enhancement of GABA release, indicating that Ca2+ influx through the 5-HT3R facilitates GABA release. Taken together, these data provide direct evidence that Ca2+ entry via presynaptic 5HT3Rs facilitates the release of GABA from hippocampal interneurons.     

Cholecystokinin releases [3H]GABA from the perfused subarachnoid space of the anaesthetized rat spinal cord

The neuropeptide cholecystokinin(26-33) (CCK) is widely distributed in the mammalian central nervous system, including the spinal cord. We have studied the possible interaction of CCK with GABA release mechanisms. Low doses of CCK-8 (1 nM) have been found to evoke calcium-dependent [3H]GABA release from an in vivo perfused spinal cord preparation in the anaesthetized rat. Tachyphylaxis was seen to the [3H]GABA releasing action of CCK-8. The injection of proglumide (150 mg/kg i.p.) totally blocked the [3H]GABA release produced by CCK-8 or by a medium containing 50 mM potassium. Substance P (10 microM) did not produce release of [3H]GABA, although in the same animals 50 mM potassium containing solutions could be shown to evoke release of [3H]GABA.     

gamma-Aminobutyric acidA (GABAA) receptors modulate [3H]GABA release from isolated neuronal growth cones in the rat

Potassium-induced release of gamma-[3H]aminobutyric acid [( 3H]GABA) from a growth cone-enriched fraction isolated from neonatal rat forebrain was inhibited by the GABA mimetic muscimol in a dose-dependent manner (IC50 15 nM). The GABA antagonist bicuculline completely reversed the effect of muscimol. Bicuculline alone slightly potentiated the K+-induced release of [3H]GABA. Baclofen, a proposed selective agonist for a bicuculline-insensitive GABAB receptor, was found to cause only a slight reduction in the K+-induced release of [3H]GABA. These results are compatible with the presence of a negative feedback mechanism mediated by GABAA receptors for controlling [3H]GABA release from growth cones of the developing rat forebrain.     

Modulation of GABA release by alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate and N-methyl-D-aspartate receptors in matrix-enriched areas of the rat striatum.

Using a new in vitro superfusion device, the release of preloaded [3H]GABA was examined in microdiscs of tissues taken from sagittal slices in matrix-enriched areas of the rat striatum. Potassium (9 mM, 15 mM) stimulated the release of [3H]GABA in a concentration- and calcium-dependent manner and the veratridine (1 microM)-evoked release of [3H]GABA was completely abolished in the presence of tetrodotoxin (1 microM). The selective glutamatergic agonist alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (1 mM) enhanced the potassium-evoked release of [3H]GABA as well as the basal outflow of [3H]GABA. This latter effect was found to be calcium-dependent, partially diminished by tetrodotoxin (1 microM), completely blocked by 6,7-dinitro-quinoxaline-2,3-dione (0.1 mM), which is generally used as an antagonist of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate receptors, but not affected by (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (MK801, 10 microM), a specific antagonist of N-methyl-D-aspartate receptors. Similarly, N-methyl-D-aspartate (1 mM) enhanced both the potassium (9 mM) and the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (1 mM)-evoked release of [3H]GABA but when used alone, due to the presence of magnesium in the superfusion medium, was ineffective on the basal efflux of [3H]GABA. A stimulatory effect of N-methyl-D-aspartate (1 mM) on the basal outflow of [3H]GABA was observed, however, when magnesium was omitted from the superfusion medium. The stimulatory effect of N-methyl-D-aspartate (1 mM) observed in the presence of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate was not potentiated by glycine (1 microM, in the presence of strychnine 1 microM) and the N-methyl-D-aspartate-evoked response seen in the absence of magnesium was not enhanced by D-serine (1 mM), suggesting that endogenous glycine is already acting on N-methyl-D-aspartate receptors. In fact, in the absence of magnesium, 7-chloro-kynurenate (1 mM) completely abolished the stimulatory effect of N-methyl-D-aspartate on the release of [3H]GABA confirming that under our conditions, the glycine site of the N-methyl-D-aspartate receptor is saturated. N-methyl-D-aspartate-evoked responses were all blocked by MK801 (10 microM). Finally, the N-methyl-D-aspartate-evoked response seen in the absence of magnesium was markedly reduced in the presence of tetrodotoxin (1 microM).(ABSTRACT TRUNCATED AT 400 WORDS)     

The effect of cochlear nerve lesion on the release of glutamate,aspartate, and GABA from cat cochlear nucleus,in vitro

Summary Pool studies of glutamate and aspartate have suggested a transmitter role for these amino acids in cochlear nerve endings. As further evidence. the K⁺-evoked release of glutamate, aspartate and GABA was measured in cat cochlear nucleus slices in vitro and compared to the release following a cochlear nerve lesion. Using [³H]glutamine as precursor, the K⁺-evoked release of glutamate and -aminobutyric acid (GABA) was respectively 4.1 and 7.2 times the spontaneous release. Using [¹⁴C]glutamate as a marker, the K⁺-evoked release of glutamate and GABA was respectively 7.1 and 2.8 times the basal release. All K⁺-evoked releases were Ca⁺⁺-dependent. Nine days after unilateral lesion of the cochlear nerve in the cat, the glutamate release decreased by about 75% on the lesioned side compared to the intact one, whereas the GABA release was not decreased. The labelled tissue glutamate, either synthesized from [³H]glutamine or labelled with [¹⁴C]glutamate, was also markedly decreased on the lesioned side. In comparison, the evoked release of aspartate, newly synthesized from [¹⁴C]glutamate, remained low and was only decreased by about 45% after cochlear nerve lesions. Comparing cat with rat cochlear nucleus, the glutamate release was similar in both animals, whereas the GABA release was much higher in the rat.It is concluded that glutamate and to a lesser extent aspartate are likely to be released from cochlear nerve terminals, supporting a transmitter role in these nerve fibres for both amino acids.     

Baclofen attenuates harmaline induced tremors in rats

Because activation of D2 dopamine receptors inhibits gamma-aminobutyric acid (GABA) release from intrapallidal nerve terminals, we measured the effects of modifiers of dopamine D2 receptors on the firing rate of single neurons in the globus pallidus (GP) of the anesthetized rat. The predominant effect of intrapallidal administration of the selective D2 agonist quinpirole was an increase in the rate of spontaneous firing while the D2 blocker sulpiride caused a decrease. The spontaneous firing of GP neurons is inhibited by stimulation of the GABAergic striatopallidal projection. We therefore measured the effects of modifiers of D2 receptors on striatal inhibition of GP neurons and found that intrapallidal quinpirole blocked the inhibitory effects of striatal stimulation while sulpiride enhanced them. These experiments show that both the spontaneous rate of firing of pallidal neurons and its modification by striatopallidal inputs is controlled by intrapallidal dopamine D2 receptors. In addition, taken together with other findings in the literature, our results suggest that activation of dopamine D2 receptors within the globus pallidus leads to inhibition of GABA release from presynaptic terminals.     

γ-Aminobutyric acid infusion in substantia nigra pars reticulata in rats inhibits ascorbate release in ipsilateral striatum

A relatively high level of extracellular ascorbate in the striatum, which is known to modulate impulse flow in striatal neurons, originates primarily from glutamate-containing corticostriatal afferents. Increasing evidence suggests that ascorbate release from these fibers is regulated by a multisynaptic loop that includes γ-aminobutyric acid (GABA) mechanisms in the substantia nigra. To assess the role that nigral GABA plays in striatal ascorbate release, extracellular ascorbate was monitored voltammetrically in the striatum during infusions of GABA into the substantia nigra pars reticulata (SNr) of awake, unrestrained rats. Compared to vehicle infusions, intranigral GABA lowered striatal ascorbate by >50%. In contrast, intranigral application of picrotoxin, a GABA antagonist, had the opposite effect. Neither GABA nor picrotoxin altered striatal 3,4-dihydroxyphenylacetic acid (DOPAC), a major dopamine metabolite. Collectively, these results indicate that intranigral GABA exerts a tonic inhibitory influence on ascorbate release in the striatum.