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Glutamate – dopamine interactions in the subthalamic nucleus. (Oral)
Boekbijdrage - Boekhoofdstuk Conferentiebijdrage
Introduction
The subthalamic nucleus (STN) is considered an important modulator in the basal ganglia. Together with the striatum, the STN forms the main input region in the motorloop circuitry [1]. It is composed of glutamatergic neurons that participate in the so called indirect pathway, projecting towards the output nuclei of the basal ganglia [substantia nigra (SN) pars reticulata and the entopeduncular nucleus] and the globus pallidus externa (GPe). It's most important inputs consist of excitatory glutamatergic projections from the cortex, GABA-ergic projections from the GPe and a direct dopaminergic input from the SN pars compacta [3,4]. Overactivity of STN neurons due to the loss of midbrain dopaminergic neurons is believed to be a key feature in Parkinson's disease. Moreover, current views of the motor circuit place the STN in a pivotal position [2]. Reciprocal interactions between dopamine (DA) and glutamate have been the subject of research, especially in the striatum and the SN pars reticulata [5]. However, little is known about these interactions in the STN.
Using in vivo microdialysis, we studied the effect of subthalamic perfusion of DA, selective D1 and D2 receptor agonists as well as NMDA on glutamate and DA release in the STN. The effect of systemic administration of L-Dopa was also evaluated. Finally, the importance of the dopaminergic innervation to the STN was evaluated by 6-hydroxydopamine (6-OHDA) lesioning of the SN.
Methods
All experiments were performed on freely moving male albino Wistar rats, weighing 280-320 g. A 1 mm microdialysis probe (MAB 4.15.1., Sweden) was placed in the left STN (AP -3.8; L -2.5; V +7.0). After collection of 8 baseline samples (15 min samples at 2.3 µL/min), pharmacological manipulation was performed. Drugs were dissolved in modified Ringer's solution (Ca2+ 2.3 mM) and administered locally via the probe for 1 hour followed by perfusion with modified Ringer's solution for 2 hours. Following compounds were perfused: dopamine (20nM), the dopamine D1 receptor agonist (SKF38393; 10-100µM), the dopamine D2 receptor agonist (quinpirole; 10-100µM) or NMDA (10-100-1000µM). NMDA-mediated effects were studied by co-perfusing NMDA with the D1 receptor antagonist SCH23390 (8µM), the D2 receptor antagonist remoxipride (4µM) or the NMDA receptor antagonist MK-801 (10µM). L-Dopa was administered systemically (25 mg/kg i.p.) with prior administration of benserazide (10 mg/kg i.p.). The effect of MK-801 (0.1 mg/kg i.p.) on the L-Dopa-induced changes in glutamate and DA was studied as described before [6]. Microdialysis samples were analysed for both glutamate and DA [7].
The 6-OHDA lesioned rats received a unilateral injection of 16 µg/4µL of 6-OHDA in the left SN 2 weeks prior to the experiment. Coordinates for the injection were AP -5.0; L -1.4; V + 8.5.
Results and Discussion
Baseline levels of DA and glutamate were 0.09 ± 0.01nM (mean ± SEM, n=6) and 116.3 ± 16.1 (mean ± SEM, n=49) respectively.
We observed that DA facilitates the release of glutamate in the STN via D1 receptors since both perfusion with DA and the D1 selective DA receptor agonist SKF38393 (100µM) significantly enhanced subthalamic glutamate levels by 230% and 260% respectively. D2 receptors are not involved since the DA D2 selective agonist quinpirole failed to influence the glutamate levels in the STN. NMDA perfusion (100µM) significantly enhanced DA release in the STN to about 170% of baseline levels. This increase coincided with an increase in extracellular glutamate levels (350%) that was D1 and D2 receptor mediated since co-perfusion with the D1 receptor antagonist SCH23390 or the D2 receptor antagonist remoxipride completely abolished the observed effects. Perfusion with MK-801 (10µM) also blocked the observed effects indicating a selective action of NMDA. Perfusion with SKF38393, remoxipride or MK-801 had no influence on extracellular glutamate levels, suggesting the absence of tonic dopaminergic and glutamatergic control on the neurotransmitter release in the STN. The NMDA mediated increase in extracellular DA and glutamate and the D1 receptor induced increase in extracellular glutamate depend on the nigral dopaminergic innervations since these effects were absent after 6-OHDA lesioning.
As expected, systemic administration of L-Dopa induced a significant increase (10-fold) in extracellular DA levels in the STN. No effect was seen on extracellular glutamate levels though. Prior administration of 0.1 mg/kg MK-801 showed a less pronounced effect on extracellular DA release after L-Dopa. Again glutamate levels were not affected.
Our work points to an important role of DA at the level of the STN. DA stimulates glutamate release in the STN via D1 receptors and NMDA-mediated release of glutamate is mediated via both D1 and D2 receptors. The dopaminergic innervations from the SN are crucial since 6-OHDA lesioning of the SN abolishes these local effects. On the other hand, DA release after systemically administered L-Dopa has little effect on subthalamic glutamate release, at least in intact rats. The result of these interactions between DA and glutamate is probably a fine tuning of the activity of subthalamic neurons, possibly critical for the coordination of motor activity.
References
1. Obeso JA, Rodriguez-Oroz M, Benitez-Temino B, Blesa FJ, Guridi J, Marin C, Rodriguez M (2008) Functional organization of the basal ganglia: therapeutic implications for Parkinson's disease. Mov. Disord. 23: S548-559.
2. Smith Y, Bevan MD, Shink E and Bolam JP (1998) Microcircuitry of the direct and indirect pathways of the basal ganglia. Neuroscience, 86: 353-387.
3. Albin RL, Young AB and Penney JB (1989) The functional anatomy of basal ganglia disorders. Trends Neurosci, 18: 366-375.
4. Parent A and Hazrati LN (1995) Functional anatomy of the basal ganglia. 2. The place of subthalamic nucleus and external pallidum in basal ganglia circuitry. Brain Res Rev, 20: 128-154.
5. Morari M, O'Connor WT, Ungerstedt U and Fuxe K (1996) Reciprocal dopamine-glutamate modulation of release in the basal ganglia. Neurochem. Int. 33: 383-397.
6. Jonkers N, Sarre S, Ebinger G, Michotte Y (2000) MK-801 influences L-DOPA induced dopamine release in intact and hemi-parkinson rats. Eur J Pharmacol, 407:281-291.
7. B. Ampe, A. Massie, J. D'Haens, G. Ebinger, Y. Michotte, S. Sarre (2007) NMDA-mediated release of glutamate and GABA in the subthalamic nucleus is mediated by dopamine: an in vivo microdialysis study in rats. J Neurochem. 103(3):1063-74.
The subthalamic nucleus (STN) is considered an important modulator in the basal ganglia. Together with the striatum, the STN forms the main input region in the motorloop circuitry [1]. It is composed of glutamatergic neurons that participate in the so called indirect pathway, projecting towards the output nuclei of the basal ganglia [substantia nigra (SN) pars reticulata and the entopeduncular nucleus] and the globus pallidus externa (GPe). It's most important inputs consist of excitatory glutamatergic projections from the cortex, GABA-ergic projections from the GPe and a direct dopaminergic input from the SN pars compacta [3,4]. Overactivity of STN neurons due to the loss of midbrain dopaminergic neurons is believed to be a key feature in Parkinson's disease. Moreover, current views of the motor circuit place the STN in a pivotal position [2]. Reciprocal interactions between dopamine (DA) and glutamate have been the subject of research, especially in the striatum and the SN pars reticulata [5]. However, little is known about these interactions in the STN.
Using in vivo microdialysis, we studied the effect of subthalamic perfusion of DA, selective D1 and D2 receptor agonists as well as NMDA on glutamate and DA release in the STN. The effect of systemic administration of L-Dopa was also evaluated. Finally, the importance of the dopaminergic innervation to the STN was evaluated by 6-hydroxydopamine (6-OHDA) lesioning of the SN.
Methods
All experiments were performed on freely moving male albino Wistar rats, weighing 280-320 g. A 1 mm microdialysis probe (MAB 4.15.1., Sweden) was placed in the left STN (AP -3.8; L -2.5; V +7.0). After collection of 8 baseline samples (15 min samples at 2.3 µL/min), pharmacological manipulation was performed. Drugs were dissolved in modified Ringer's solution (Ca2+ 2.3 mM) and administered locally via the probe for 1 hour followed by perfusion with modified Ringer's solution for 2 hours. Following compounds were perfused: dopamine (20nM), the dopamine D1 receptor agonist (SKF38393; 10-100µM), the dopamine D2 receptor agonist (quinpirole; 10-100µM) or NMDA (10-100-1000µM). NMDA-mediated effects were studied by co-perfusing NMDA with the D1 receptor antagonist SCH23390 (8µM), the D2 receptor antagonist remoxipride (4µM) or the NMDA receptor antagonist MK-801 (10µM). L-Dopa was administered systemically (25 mg/kg i.p.) with prior administration of benserazide (10 mg/kg i.p.). The effect of MK-801 (0.1 mg/kg i.p.) on the L-Dopa-induced changes in glutamate and DA was studied as described before [6]. Microdialysis samples were analysed for both glutamate and DA [7].
The 6-OHDA lesioned rats received a unilateral injection of 16 µg/4µL of 6-OHDA in the left SN 2 weeks prior to the experiment. Coordinates for the injection were AP -5.0; L -1.4; V + 8.5.
Results and Discussion
Baseline levels of DA and glutamate were 0.09 ± 0.01nM (mean ± SEM, n=6) and 116.3 ± 16.1 (mean ± SEM, n=49) respectively.
We observed that DA facilitates the release of glutamate in the STN via D1 receptors since both perfusion with DA and the D1 selective DA receptor agonist SKF38393 (100µM) significantly enhanced subthalamic glutamate levels by 230% and 260% respectively. D2 receptors are not involved since the DA D2 selective agonist quinpirole failed to influence the glutamate levels in the STN. NMDA perfusion (100µM) significantly enhanced DA release in the STN to about 170% of baseline levels. This increase coincided with an increase in extracellular glutamate levels (350%) that was D1 and D2 receptor mediated since co-perfusion with the D1 receptor antagonist SCH23390 or the D2 receptor antagonist remoxipride completely abolished the observed effects. Perfusion with MK-801 (10µM) also blocked the observed effects indicating a selective action of NMDA. Perfusion with SKF38393, remoxipride or MK-801 had no influence on extracellular glutamate levels, suggesting the absence of tonic dopaminergic and glutamatergic control on the neurotransmitter release in the STN. The NMDA mediated increase in extracellular DA and glutamate and the D1 receptor induced increase in extracellular glutamate depend on the nigral dopaminergic innervations since these effects were absent after 6-OHDA lesioning.
As expected, systemic administration of L-Dopa induced a significant increase (10-fold) in extracellular DA levels in the STN. No effect was seen on extracellular glutamate levels though. Prior administration of 0.1 mg/kg MK-801 showed a less pronounced effect on extracellular DA release after L-Dopa. Again glutamate levels were not affected.
Our work points to an important role of DA at the level of the STN. DA stimulates glutamate release in the STN via D1 receptors and NMDA-mediated release of glutamate is mediated via both D1 and D2 receptors. The dopaminergic innervations from the SN are crucial since 6-OHDA lesioning of the SN abolishes these local effects. On the other hand, DA release after systemically administered L-Dopa has little effect on subthalamic glutamate release, at least in intact rats. The result of these interactions between DA and glutamate is probably a fine tuning of the activity of subthalamic neurons, possibly critical for the coordination of motor activity.
References
1. Obeso JA, Rodriguez-Oroz M, Benitez-Temino B, Blesa FJ, Guridi J, Marin C, Rodriguez M (2008) Functional organization of the basal ganglia: therapeutic implications for Parkinson's disease. Mov. Disord. 23: S548-559.
2. Smith Y, Bevan MD, Shink E and Bolam JP (1998) Microcircuitry of the direct and indirect pathways of the basal ganglia. Neuroscience, 86: 353-387.
3. Albin RL, Young AB and Penney JB (1989) The functional anatomy of basal ganglia disorders. Trends Neurosci, 18: 366-375.
4. Parent A and Hazrati LN (1995) Functional anatomy of the basal ganglia. 2. The place of subthalamic nucleus and external pallidum in basal ganglia circuitry. Brain Res Rev, 20: 128-154.
5. Morari M, O'Connor WT, Ungerstedt U and Fuxe K (1996) Reciprocal dopamine-glutamate modulation of release in the basal ganglia. Neurochem. Int. 33: 383-397.
6. Jonkers N, Sarre S, Ebinger G, Michotte Y (2000) MK-801 influences L-DOPA induced dopamine release in intact and hemi-parkinson rats. Eur J Pharmacol, 407:281-291.
7. B. Ampe, A. Massie, J. D'Haens, G. Ebinger, Y. Michotte, S. Sarre (2007) NMDA-mediated release of glutamate and GABA in the subthalamic nucleus is mediated by dopamine: an in vivo microdialysis study in rats. J Neurochem. 103(3):1063-74.
Boek: Proceedings of the 13th International Conference on In Vivo Methods
Series: Monitoring Molecules in Neuroscience
Pagina's: 75-77
Aantal pagina's: 3
Jaar van publicatie:2010
Trefwoorden:subthalamic nucleus, microdialysis, basal ganglia, Parkinson's disease, L-DOPA, 6-OHDA rat model