Summary

Maximilian Darrah, John Partridge, Stefano Vicini

Department of Biology, Department of Pharmacology and PhysiologyGeorgetown University, Washington, D.C.GABAergic control of excitatory input to striatal spiny projection neurons (SPNs)

Motor Function Basal Ganglia

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Striatonigral and Striatopallidal pathwaysGerfen, (2006) Nat Neurosci 9(2):157-8

(D2+)(D1+)These outputs facilitates or inhibits movement initiation and control

3MSNs are GABAergic projections neurons and make up 95% of cells in the striatum and belong to one of two pathways. Those in the direct pathway express dopamine D1 receptors and the neuropeptide substance P. They project directly to the SNr and are thought to facilitate movement whereas those in the indirect pathway express D2 Rs and the neuropeptide enkephalin. Their output gets to the SNr indirectly via a series of relay neurons on the GPe and STN. These are thought to facilitate neurons.

Striatal Innervation

GlutamateGABAGABA-Excitatory inputs from cortex, thalamus-Inhibitory inputs from GABAergic interneurons and SPNs/MSNs collaterals-Modulatory inputs (dopamine) from SNcAdapted from Tepper et al., 2004 Curr Opin Neurobiol 14:685.

LTS

TH

SPN

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Postsynaptic GABA current

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Subunit composition determines pharmacology and function of the receptor

pharmacological sensitivity channel kineticsdesensitization ratesregional and subcellular localization.

Tija et al. 2008 Nature Reviews Neuroscience 9:3311-6, 1-3, 1-3, , , 1-3, , 1-3

Neil Harrison: Navigating Neuronal Pathways

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MethodsCalcium ImagingRecordings at room temperatureStimulation with a bipolar Tungsten electrodeDrug applied viaytubing system

Acute slice prepPost-natal day 13-21Dissect out brainCut 250m coronal slices containing the striatumIncubated in artificial CSF

Y tubingStimulating Electrode

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GCaMP3 Protein

Is a genetically encoded calcium indicator (GECI)Consists of Green Fluorescent Protein, Calmodulin, and M13Modulates fluorescence based on presence or absence of calcium.Cell specificity: drd2;GCaMP3 and drd1;GCaMP3

Figure A taken from Lindenburg et al ChemBioChem(13): 349-351

[Ca2+]iSpikes

50% F/F0 20 mV

10 s

60xm24 Area 512x512 Mag 60xFS 150 Laser 4.1 GainA 40 Cuml 6 PH 200 Frames 400 time 1:42 Rate 2.45 hz Temperature: 23.3 CDHPG 20 uM at 20 wash 60NMDA 20 uM at 100 wash 183DHPG 20 uM 253 wash 295 (1 min after NMDA)KCl at 360 w 380Start timer with addition of NMDAGood, many cells respond

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Synaptic Activation of Striatopallidal SPNs in a drd2;GCaMP3 Mouse

Movie at 15x speed

GABAzine 10 M3.5 s100% df/f13456279

350 ms25 % df/f

GABAzineWashControlGabazine effects in drd2 SPNs

2060100

Cell1

Cell2

Cell3

Cell4

Cell5

Cell6

Cell7

Cell8

Cell9Peak (dF/F0)PeakDecay time

Latency350 msT of PeakGabazine affects peak fluorescence in drd2 SPNs

Gabazine effects are specific for drd2 SPNs

0%50%100%150%200%P13-P15

drd1

drd2P19-21**p< 0.005 to control% control*

Diazepam effects

0%50%100%150%

P13-P15P19-P21

drd1

drd2*% control

Conclusions and Future DirectionsSynaptic excitation of SPNs evokes calcium transients that are variable within and between cells.

Comparing Gabazines effects on SPNs in drd1;GCaMP3 and drd2;GCaMP3 mice, we revealed GABAergic control of the indirect pathway SPNs is much stronger than that of the direct pathway.

These data are supported by the inhibitory effect of diazepam in the drd2 but not drd1 SPNs and are relevant for the understanding and treatment of movement disorders.

Special Thanks to the Vicini LabStefano ViciniJohn PartridgeCarissa WinlandNour Al-Muhtasib Supported by the Howard Hughes Medical Institute Scholars Program

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