Dialysate Levels Of Glutamate Research Articles

Striatal and nigral muscarinic type 1 and type 4 receptors modulate levodopa-induced dyskinesia and striato-nigral pathway activation in 6-hydroxydopamine hemilesioned rats

Acetylcholine muscarinic receptors (mAChRs) contribute to both the facilitation and inhibition of levodopa-induced dyskinesia operated by striatal cholinergic interneurons, although the receptor subtypes involved remain elusive. Cholinergic afferents from the midbrain also innervate the substantia nigra reticulata, although the role of nigral mAChRs in levodopa-induced dyskinesia is unknown. Here, we investigate whether striatal and nigral M1 and/or M4 mAChRs modulate dyskinesia and the underlying striato-nigral GABAergic pathway activation in 6-hydroxydopamine hemilesioned rats. Reverse microdialysis allowed to deliver the mAChR antagonists telenzepine (M1 subtype preferring), PD-102807 and tropicamide (M4 subtype preferring), as well as the selective M4 mAChR positive allosteric modulator VU0152100 in striatum or substantia nigra, while levodopa was administered systemically. Dyskinetic movements were monitored along with nigral GABA (and glutamate) and striatal glutamate dialysate levels, taken as neurochemical correlates of striato-nigral pathway and cortico-basal ganglia-thalamo-cortical loop activation. We observed that intrastriatal telenzepine, PD-102807 and tropicamide alleviated dyskinesia and inhibited nigral GABA and striatal glutamate release. This was partially replicated by intrastriatal VU0152100. The M2 subtype preferring antagonist AFDX-116, used to elevate striatal acetylcholine levels, blocked the behavioral and neurochemical effects of PD-102807. Intranigral VU0152100 prevented levodopa-induced dyskinesia and its neurochemical correlates whereas PD-102807 was ineffective. These results suggest that striatal, likely postsynaptic, M1 mAChRs facilitate dyskinesia and striato-nigral pathway activation in vivo. Conversely, striatal M4 mAChRs can both facilitate and inhibit dyskinesia, possibly depending on their localization. Potentiation of striatal and nigral M4 mAChR transmission leads to powerful multilevel inhibition of striato-nigral pathway and attenuation of dyskinesia.

Investigation of the Role of AT2 Receptors in the Nucleus Tractus Solitarii of Normotensive Rats in Blood Pressure Control.

AimThe nucleus tractus solitarii (NTS) densely expresses angiotensin II type 2 receptors (AT2R), which are mainly located on inhibitory gamma-aminobutyric acid (GABA) neurons. Central AT2R stimulation reduces blood pressure, and AT2R stimulation in the rostral ventrolateral medulla (RVLM), mediates a hypotensive response through a GABAergic mechanism. We aimed to test the hypothesis that an AT2R mediated inhibition of the GABA release within the NTS might be involved in this hypotensive response, by assessing possible alterations in blood pressure and heart rate, as well as in GABA levels in normotensive Wistar rats.MethodsIn vivo microdialysis was used for measurement of extracellular GABA levels and for perfusion of the selective AT2R agonist, Compound 21, within the NTS. Our set-up allowed to determine simultaneously the excitatory glutamate dialysate levels. The mean arterial pressure and heart rate responses were monitored with a pressure transducer.ResultsLocal perfusion of Compound 21 into the NTS did not modify blood pressure and heart rate, nor glutamate and GABA levels compared to baseline concentrations. A putative effect was also not unmasked by concomitant angiotensin II type 1 receptor blockade with candesartan. Positive control experiments confirmed that the experimental set up had enough sensitivity to detect a reduction in GABA dialysate levels and blood pressure.ConclusionThe results did not provide evidence for a role of the AT2R within the NTS in the control of blood pressure, nor for an interaction with local GABAergic signaling in normotensive rats.

Neuropeptide Y increases in vivo hippocampal extracellular glutamate levels through Y1 receptor activation

Neuropeptide Y's (NPY) anticonvulsant effect is generally attributed to its inhibitory effect on glutamate release from presynaptic nerve terminals, which is nicely demonstrated in in vitro settings. To date no study has attempted to investigate the effect of NPY in vivo on extracellular (EC) glutamate levels thus, via intracerebral microdialysis, we determined NPY's effect on hippocampal glutamate concentrations in vivo, and consequently the involvement of Y1 receptors to this effect. NPY or the Y1 agonist D-His26-NPY was intrahippocampally administered in rats for 2h, during which the hippocampal glutamate dialysate levels were monitored. Pilocarpine was subsequently co-administered with NPY or D-His26-NPY to determine their effect on pilocarpine-induced limbic seizures. Unexpectedly we noted that intrahippocampal administration of NPY or D-His26-NPY increased glutamate dialysate levels in a reproducible manner. NPY attenuated pilocarpine induced seizures, whereas D-His26-NPY did not. To clarify the role of Y1 receptors in NPY's glutamatergic effect, NPY was co-administered with the selective Y1 antagonist BVD10. Hippocampal Y1 receptor blockade prevented the NPY-induced increase in hippocampal glutamate, proving that this induced glutamate increase is clearly Y1 receptor mediated. This is the first evidence that NPY enhances hippocampal EC glutamate overflow in vivo via hippocampal Y1 receptors without interfering with or contributing to NPY's anticonvulsant effect. Whilst this finding contrasts with the supposed glutamatergic hypothesis for NPY in the hippocampus, it is of significance to further assist in deciphering NPY's mechanisms of action in in vivo settings.

Influence of glial cells in the dopamine releasing effect resulting from the stimulation of striatal δ-opioid receptors

Recent data indicate that striatal dopamine release induced by stimulation of δ-opioid receptors is a consequence of glutamate release. However, glial cells, which mainly support glutamate uptake and are involved in glutamate signaling and potentially express δ-opioid receptors, could participate to this effect. The present study investigates the contribution of glial cells in the releasing effects of [ d-Pen2, d-Pen5]-enkephalin (DPDPE) by using the gliotoxin l-α-aminoadipate ( l-αAA). Initially, we evaluated the early influence of l-αAA local infusion (10 μg/μL) on dialysate levels of glutamate and dopamine under basal or DPDPE treatment conditions. l-αAA produced a significant increase of glutamate and dopamine in dialysates (+76% and +50% respectively) and the concomitant infusion of DPDPE (10 μM) significantly enhanced this effect in an additive manner (+110% and +44% respectively). Secondly, we assessed the DPDPE effects on striatal glutamate and dopamine dialysate levels, 2 days after an intra-striatal injection of l-αAA which produced destruction of glial cells. This lesion, decreasing the basal glutamate dialysate level as well as its tissue content (by 55% and 36% respectively), prevented the increase in glutamate and dopamine extracellular levels induced by DPDPE. This result confirmed that the DPDPE-induced dopamine release requires an initial glutamate release. However, this effect could reflect a major disruption of glutamatergic transmission caused by the toxin, as suggested by the local infusion of glutamine (2.5 mM) which, in lesioned rats, prevented the decrease in the basal extracellular content of glutamate and restored the DPDPE-induced increase in glutamate and dopamine dialysate levels. Therefore, these results indicate that, although glial cells are essential to maintain functional glutamatergic neurotransmission, they are not directly involved in the process by which stimulation of striatal δ-opioid receptors induces extracellular glutamate release and, consecutively, dopamine release.

Impaired formalin-evoked changes of spinal amino acid levels in diabetic rats

To investigate mechanisms by which diabetes alters sensory processing, we measured levels of amino acid neurotransmitters in spinal dialysates from awake, unrestrained control and diabetic rats under resting conditions and following hind paw formalin injection. Under resting conditions, glutamate concentrations in spinal dialysates were significantly ( P < 0.05) decreased in diabetic rats compared to those of control rats whereas aspartate, taurine, glycine and citrulline remained unchanged and GABA was significantly ( P < 0.05) increased. Noxious stimulation of the hind paw by subcutaneous injection of 0.5% formalin into the dorsum caused a defined flinching behavior in the afflicted paw, and the amount of flinching was significantly ( P < 0.05) greater in diabetic rats than in controls. Paw formalin injection significantly ( P < 0.05) increased dialysate levels of glutamate, aspartate, taurine, glycine and citrulline by 3- to 4-fold above basal in both control and diabetic rats. The concentration of glutamate in dialysate samples collected immediately after paw formalin injection remained significantly ( P < 0.05) lower in diabetic rats compared to those in controls. Formalin injection did not alter dialysate GABA concentrations in control rats, whereas in diabetic rats there was an increase of 151 ± 15% above basal levels. These findings indicate that the selective depression of basal and stimulus-evoked glutamate levels in the spinal cord of diabetic rats occurs in parallel with elevated spinal GABA levels. Because increased pain-associated behavior is accompanied by an attenuated spinal glutamate spike following paw formalin injection, hyperalgesia in diabetic rats does not appear to be secondary to enhanced glutamatergic input to the spinal cord.

Microdialysis of the lateral and medial temporal lobe during temporal lobe epilepsy surgery

Background This study was undertaken to establish whether, in temporal lobe epilepsy (TLE), there are relative differences between the lateral and ipsilateral medial temporal lobe in the extracellular levels of 3 of the human brain's major neuroactive amino acids. Methods Seven generally anesthetized patients with TLE undergoing anatomically standardized resective surgery had at operation microdialysis catheters inserted within the middle temporal gyrus (ie, lateral temporal lobe) and anterior hippocampus (ie, medial temporal lobe). Surface electrocorticography (ECoG) recordings were also obtained. Samples of 10-minute dialysate were quantified for glutamate, aspartate, and gamma-aminobutyric acid (GABA) using high-performance liquid chromatography; corresponding ECoG data were assessed for epileptiform activity. Where available, resection tissue was subjected to histopathological analysis. Results The ratio of mean “sample 3” dialysate levels of glutamate, aspartate, and GABA was approximately 20:2:1, respectively, in both the minimally epileptiform lateral (n = 7) and medial (n = 5) temporal lobe; between the 2 sets of samples, these levels were not significantly different ( P > 0.05 for each amino acid studied). From the vigorously epileptiform medial temporal lobe of 2 patients, sample 3 dialysate levels of the excitatory amino acids glutamate and aspartate were found in considerably greater concentrations (between 15- and 37-fold) with correspondingly less dramatic increases of the inhibitory amino acid GABA (more than 11- and 13-fold). Laterally resected tissue (obtained in 3 cases) did not demonstrate significant cortical or subcortical abnormalities; medial resection tissue from all patients demonstrated, in varying degrees, hippocampal sclerosis. Conclusions In the absence of significant tissue hyperexcitability, despite known differences in local cellular and/or histopathological architecture, the extracellular relationship among glutamate, aspartate, and GABA is not dissimilar in both the lateral and ipsilateral medial temporal lobe of TLE patients. Considerable disparity in dialysate levels recovered (eg, from the vigorously epileptiform medial temporal lobe) may be related to the functional (ie, hyperexcitable) status of the sampled tissue.

Involvement of corticostriatal glutamatergic terminals in striatal dopamine release elicited by stimulation of δ‐opioid receptors

We have previously shown that striatal dopamine release induced locally by a delta-opioid receptor agonist was totally inhibited by a glutamate N-methyl-D-aspartate receptor antagonist, indicating the involvement of glutamatergic receptors in this effect. The aim of the present study was to specify this mechanism. Firstly, we investigated the effect of [D-Pen2,D-Pen5]-enkephalin (DPDPE) on glutamate release in rats by intrastriatal microdialysis. The infusion of DPDPE (10 microm) enhanced the glutamate content in dialysate by approximately 34%, an effect which did not appear to result from inhibition of glutamate uptake. We then considered the consequences of a unilateral thermocoagulation of the frontal cortex on either glutamate or dopamine release induced by stimulation of delta-opioid receptors 2 days later. This lesion, which decreased the glutamate content in ipsilateral striatum by approximately 30%, totally prevented the increase in dialysate levels of glutamate induced by DPDPE. Moreover, whereas DPDPE (10 microm) was found to increase the striatal dopamine release in intact animals by approximately 59%, this effect was also completely suppressed by the cortical lesion. Finally, we studied the effect of the lesion on the [3H]-DPDPE binding to striatal membranes prepared from the whole striatum. In the ipsilateral striatum a significant decrease in this [3H]-DPDPE binding (by approximately 18%) was found 2 days after the lesion. Our results indicate that the increase in striatal dopamine release induced by DPDPE probably depends on glutamate release from corticostriatal glutamatergic afferents in response to the stimulation of delta-opioid receptors located on terminals of these neurons.

A dual probe characterization of dialysate amino acid levels in the medial prefrontal cortex and ventral tegmental area of the awake freely moving rat

Dual probe microdialysis was employed to characterize the origins of dialysate glutamate, aspartate and gamma-aminobutyric acid (GABA) in the medial prefrontal cortex (mPfc) and to investigate functional interactions between the mPfc and ventral tegmental area (VTA) in awake, freely moving rats. Perfusion with elevated potassium (K +; KCl, 100 mM, 20 min), low Ca 2+ (0.1 mM, 60 min) or tetrodotoxin (TTX, 10 μM, 100 min) was performed in the mPfc and dialysate levels of glutamate, aspartate and GABA were measured locally and in the VTA. Elevated K + in the mPfc rapidly increased dialysate glutamate and aspartate locally (+90±10 and +41±9% from basal, respectively) and in the VTA (+71±14 and +42±14%, respectively). MPfc GABA was also rapidly increased (+241±62%) while VTA GABA was not affected. Perfusion with low Ca 2+ in the mPfc decreased local glutamate, aspartate and GABA (−26±8; −35±7 and −45±8%, respectively) and decreased only GABA (−40±5%) in the VTA. Intra-mPfc TTX increased glutamate and aspartate locally (+82±23 and +54±27%, respectively) and in the VTA (+84±18 and +38±17%, respectively). In contrast, intra-mPfc TTX decreased local GABA (−33+6%) while VTA GABA levels were not affected. Taken together, these data confirm the influence of the mPfc upon the ipsilateral VTA and provide evidence for two neuronal pools which contribute to basal extracellular mPfc and VTA glutamate, aspartate and GABA levels, the first pool derived from Na +- and Ca 2+-dependent release and the second derived from voltage-dependent reuptake.

Extracellular glutamate and other metabolites in and around RG2 rat glioma: an intracerebral microdialysis study.

The current study determined the extracellular content of glutamate, 10 additional amino acids, lactate, glucose and some antioxidants in a rodent model of malignant glioma, its peritumoral space and the adjacent cortex. RG2 tumors were induced in the right frontal cortex of Fischer-344 rats (n = 10) by a standardized procedure to obtain a maximum sagittal tumor width of 3-4 mm diameter. After confirmation of tumor growth and localization by contrast enhanced MRI three microdialysis probes were implanted simultaneously in the cortex: at the tumor implantation site (tumor), 2 mm caudally, brain around tumor (BAT) and 4 mm caudally (cortex) to the site of implantation. Dialysate concentrations of glutamate were increased 3.9-fold in tumor and 2-fold in BAT compared with cortex. Glycine was elevated 11.4-fold in tumor and 2.6-fold in BAT. Lactate was increased 1.7-fold in tumor, 1.2-fold in BAT. Levels of glucose, ascorbic acid and uric acid were not significantly different in tumor, BAT and cortex. The increased dialysate levels of glutamate and glycine in the peritumoral space may contribute to impaired neuronal function and epileptiform activity associated with this tumor type in humans.

GABA and glutamate levels in the substantia nigra reticulata following repetitive cerebral ischemia in gerbils.

Repetitive cerebral ischemia produces more severe damage than a similar single duration insult. We have previously shown that, in gerbils, damage in the substantia nigra reticulata (SNr) is seen with repetitive insults rather than a single insult. We have also shown that there is a progressive decrease in the extracellular GABA in the striatum in the days preceding such damage, speculating that a loss of GABA may be in part responsible for this damage. This study evaluates the GABA levels in the SNr in animals exposed to repetitive ischemic insults. Each animal received a total of three ischemic insults of 3-min duration at hourly intervals. In vivo microdialysis was carried out to analyze the GABA and glutamate dialysate levels on Days 1, 3, 5, 7, and 14 following the ischemic insult. In the control and treated (ischemic) animals, there was a significant increase in the GABA levels with the introduction of nipecotic acid on Days 1, 3, 5, and 14. However, on Day 7 there was a significant attenuation in the GABA response to nipecotic acid in the treated animals in comparison to the controls. The glutamate levels in the treated animals were similar to the control animals on Days 1, 3, 5, and 7. However, on Day 14 the glutamate levels were significantly lower than on previous days. Our experiments for the first time measure extracellular glutamate and GABA responses in the SNr in animals exposed to repetitive ischemic insults. Our experiments show that there is a significant decrease in the GABA concentrations at a time when ischemic damage is developing in this region. This confirms our hypothesis that a decrease in GABA may be one factor contributing to neuronal damage during the period following repetitive ischemic insults. Further, the rebound increase in GABA levels on Day 14 with a concomitant fall in glutamate levels would indicate that reparative processes are still active in the 2 weeks following the insult.

Evidence against high extracellular glutamate promoting the elicitation of spreading depression by potassium.

This study ascertains whether high extracellular glutamate contributes to the initiation of spreading depression (SD) by K+. Two microdialysis probes, each incorporating an electrode to record the extracellular direct current (DC) potential at the elicitation site, were implanted symmetrically in the cortex of anesthetized rats. Recurrent SD was triggered by perfusion of 130 mM K+ through the microdialysis probe for 20 min. On one side, this medium was supplemented with increasing concentrations of glutamate (0.1-1 mM) or of the selective glutamate uptake inhibitor 1-trans-pyrrolidine-2,4-dicarboxylate (L-trans-PDC: 1-10 mM). The effects of L-trans-PDC on extracellular glutamate and basal DC potential were studied in separate experiments. Application of K+ for 20 min consistently elicited five to seven waves of SD. Increasing the concentration of glutamate in the perfusion medium did not alter SD elicitation. Application of L-trans-PDC concentration dependently increased the dialysate levels of glutamate (by approximately 19-fold with 10 mM L-trans-PDC) but, unexpectedly, reduced SD elicitation. These data do not support the hypothesis that SD is elicited because high extracellular glutamate resulting from exocytosis and/or reversal of glutamate uptake depolarizes adjacent neurons. As SD elicitation requires activation of N-methyl-D-aspartate (NMDA) receptors, these results also illustrate that sensitivity of a pathological or experimental event to NMDA receptor antagonists does not necessarily imply involvement of increased extracellular glutamate. This does not rule out a selective action of glutamate, transiently released from presynaptic vesicles, on immediately juxtaposed postsynaptic receptors.

Effects of methylprednisolone on extracellular lactic acidosis and amino acids after severe compression injury of rat spinal cord.

We evaluated in rats with severe spinal cord compression at T8-9 the influence of methylprednisolone (MP) on lactic acidosis and extracellular amino acids, which may cause secondary, perifocal injuries of the cord. MP (30 mg/kg) was given intravenously 30 min before compression and hourly thereafter (15 mg/kg). Other rats with compression, given saline, served as controls. Samples from the extracellular fluid of one dorsal hom were collected by microdialysis and analyzed by HPLC. Microdialysis was performed for 1.5 h to establish basal levels. Samples were collected for 3 h after compression. MP-treated rats showed a reduction of dialysate lactic acid and arginine levels during the first 1-2 h after trauma. The mean dialysate levels of glutamate in MP-treated rats were lower than those of the controls, but the difference was not statistically significant. MP treatment did not influence dialysate levels of aspartate, glutamine, histidine, glycine, threonine, taurine, alanine, GABA, and tyrosine. Our study shows that MP has several effects, including reduced lactic acid formation, reduced levels of arginine (the substrate for nitric oxide production), and a trend toward decreased extracellular accumulation of the excitotoxic amino acid glutamate. We conclude that MP has the capacity to change the composition of the extracellular edema fluid after trauma to the spinal cord. These changes may counteract free radical formation and may be important mechanisms by which MP exerts its beneficial actions.

Microdialysis as a tool to measure dietary and regional effects on the complete profile of extracellular amino acids in the hypothalamus of rats

Regional and dietary-induced changes in hypothalamic extracellular amino acid concentrations were examined. Microdialysis probes were simultaneously implanted in the hypothalamic paraventricular nucleus (PVN) and the lateral hypothalamus (LH) of anesthetized rats and perfused at a rate of 2 μl/min. Dialysates were collected every 20 min for l h prior to gavage of a balanced amino acid mixture (0.85 g patterned after 1 g of chicken egg albumin) and then every 20 min for 3 h after treatment. Tail vein blood samples were also collected. Marked changes in plasma levels of most amino acids were evident immediately following the amino acid gavage. In the PVN, concentrations of isoleucine, leucine, methionine and valine all increased within 40 min, whereas significant decreases in glutamine, histidine and taurine were observed in the LH. In a separate study, PVN extracellular amino acid concentrations were examined in awake, freely-behaving rats following gavage of equicaloric loads of a balanced amino acid mixture, glucose (0.89 g) or water. Dialysate levels of glutamate, isoleucine, leucine, methionine, threonine, tyrosine and valine showed reliable increases after amino acid treatment, although the overall time course of these effects differed somewhat. The amino acid profile of the PVN was, in general, unaffected by glucose administration. These findings suggest that specific brain regions may respond uniquely to amino acid ingestion and further imply that dietary composition may influence the amino acid profiles of the extracellular fluid in brain.

Dopamine D 1 and D 2 receptor antagonism differentially modulates stimulation of striatal neurotransmitter levels by [formula omitted] acid

The effects of local perfusion with the dopamine D 1 receptor antagonist SCH 23390 and the dopamine D 2 receptor antagonist raclopride on basal and N- methyl- D - aspartate (NMDA) stimulated dopamine, γ-aminobutyric acid (GABA) and glutamate levels in the dorsolateral striatum were monitored using in vivo microdialysis anaesthetized rat. Local perfusion (90 min) with SCH 23390 and raclopride (1 and 10 μM) dose dependently increased basal striatal dopamine release whereas GABA and glutamate dialysate levels were unaffected. Local perfusion (10 min) with the 1 mM concentration of NMDA increased striatal dopamine, GABA and glutamate outflow. However, when perfused together with SCH 23390 (1 μM) NMDA inhibited glutamate efflux. Moreover, in the presence of SCH 23390 (10 μM) the NMDA induced rise in dopamine and GABA levels was partly prevented. On the other hand, raclopride 1 μM enhanced the NMDA stimulated GABA efflux while at 10 μM it partly counteracted the NMDA induced dopamine release. These data demonstrate that NMDA induced changes in striatal neurotransmitter outflow are effectively modified by dopamine D 1 and D 2 receptor blockade.

Amino acid neurotransmitters in nucleus tractus solitarius: an in vivo microdialysis study.

Amino acid neurotransmitters in the nucleus tractus solitarius (NTS) are thought to play a key role in the mediation of visceral reflexes and glutamate has been proposed as the neurotransmitter of visceral afferent nerves projecting to this region. The present studies sought to characterize the use of in vivo microdialysis to examine extracellular fluid levels of amino acids in the NTS of anesthetized rats. Using a microdialysis probe that was 450 microns in length and a sensitive HPLC assay for amino acids, amino acids could be measured in dialysate samples collected from the NTS. Perfusion of the microdialysis probe with 60 mM K+, to elicit depolarization of nerve terminals in the vicinity of the probe, resulted in increased dialysate fluid levels of aspartate, glutamate, glycine, taurine, and GABA. In contrast, glutamine and tyrosine were decreased and other amino acids were not significantly affected. Prior removal of the ipsilateral nodose ganglion did not alter the K(+)-evoked changes in dialysate levels of any of these amino acids. Electrical stimulation of the vagus nerves, using a variety of stimulus parameters, did not significantly alter dialysate levels of glutamate or any of the other amino acids that were measured. Blockade of glutamate uptake with dihydrokainate increased dialysate levels of glutamate, aspartate, and GABA, but in the presence of dihydrokainate vagal stimulation did not alter dialysate levels of these amino acids. The results show that in vivo microdialysis can be used to examine amino acid efflux in the rat NTS and provide further evidence for amino acidergic neural transmission in the NTS. However, these studies fail to support the hypothesis that vagal afferents release glutamate or aspartate.