Suppression of seizure-like activity by optogenetic stimulation in reticular

thalamus

Student: Hsueh Tai-En 薛岱恩 Department of Psychology, NTU

Advisors: Dr. Shyu Bai-Chuang 徐百川 老師 Chang Wei-Jen 張瑋仁 , Chang Wei-Pang 張維邦 IBMS, Academia Sinica

Importance of seizure• Seizure is abnormal excessive or synchronous neural activity in the

brain, producing a physical convulsion, minor physical signs, thought disturbances, or a combination of symptoms (Fisher et al., 2005).

• Seizure affects 1 % of population, and 30 % are suffered from drug-resistant epilepsy. Clinical application uses non-invasive approaches or deep brain stimulation for the treatment of drug-resistant seizures.

• Thalamus plays an important role on modulating the brain cortical seizure activity (Chang et al., 2011;Polack 2009; Ates et al., 2005; Blumenfeld, 2002).

Wei-Pang Chang(2011). Epilepsia, 52(12)

Thalamocortical mechanism of seizure

Jon-Paul(2003). TRENDS in Pharmacological Sciences, Vol.24, No.10

Red: inhibitory GABA-containing neuron

Green: excitatory glutamate-containing neuron

Reticular Thalamus Nucleus

GABA

Relation between Seizure and reticular thalamus

• A gain in GABAergic inhibitory strength in thalamus reduces the length and power of absence seizure (Schofield et al., 2009).

• The net effect of increased GABAergic synaptic transmission within NRT reduces synchronized burst-firing of thalamic relay neurons. GABAA receptor-mediated inhibition of NRT outflow decrease the likelihood of absence seizures ( Hosford et al., 1997).

GABAergicinterneuron

GABA

Seizure

The problem• Pharmacology• Electrical stimulation

Using light stimulation

Optogenetic• Optogenetic technology combines genetic targeting of specific

neurons or proteins with optical technology for imaging or control of the targets within intact, living neural circuits (Deisseroth et al., 2006).

Aravanis. J. Neural Eng,4. 2007

Pastrana. Nature Methods 8. 2011

Channelrhodopsins

• Hypothesis: Activation of nRT will modulate cortical seizure• Aim: 1. Use Pv-ChR2 mice as model, and verify the expression of channelrhodopsin. 2. Induce cortical seizure-like activity by injecting pentylenetetrazol

(PTZ) (GABAA antagonist).

3. Use light to specifically activate nRT neuron output to investigate the effect of nRT activation during seizure.

Material & Method• Pv-chr2 transgenic mice

• Michigan probe

- In the reticular thalamic nucleus, nearly all neurons are parvalbumin-immunoreactive (Csillik et al., 2005).

16 electrodelaser

Tungsten electrode corpus callosum (CC) stimulate

Michigan probeRecording: Primary somatosensory cortex

Optic fiber-optic electrodeReticular thalamus stimulate

pv-chr2 transgenic mice Injected 60mg/kg PTZ

Experiment setup

CC

S1

RT

Procedure

Record 30 minsLight stimulate

+Induce seizure

Turn on lightInduce seizure6 times

Light stimulationCC stimulation

Anti EYFP Anti PV merge100 mm

2 mm

The expression of channelrhodopsin

cerebellum

Reticular thalamus

Olfactory bulb

HippocampusCortex

Light stimulation modulates RT and thalamus firing rate

Stimulation reticular thalamus(RT)

Stimulation on thalamus(Th)

Cx Th

Light stimulation in RT changes the firing pattern in cortex

Primary somatosensory cortex(S1)

P 1.82 mmL 2.5mm

Reticular thalamus (RT)

P 3.5mmL 2.0mm

Corpus callosum(CC) stimulation induces seizureSuperficial layer

0.5 mV

300 ms

Deeper layerCorpus callosum stimulate

Control

Antiepleptic drug Ethoxusimide blocks CC induced seizure

2 mV

1 sec

CC stimulation

Control

light stimulation 5Hz pulse duration 100ms duration 20s

No light stimCC stim 10HZ 500uA 4s

Light stimulation suppressing CC induced seizure

2 mV

1 sec

control 5Hz 20s 10Hz 20s 20HZ 20s 50Hz 20s 100Hz 20s 10Hz 30s 20HZ 30s 50Hz 30s 100Hz 30s cont. 1s cont. 10s cont. 20s0

10

20

30

40

50

60

70

80

90

100

succ

ess i

nduc

e ra

te(%

)

Most effective light stimulation pattern in suppressing seizure is 100Hz 30s light pulse

Different light stimulation condition in suppressing seizure

*

Light stimulation in RT is necessary for suppressing seizure

>60%30%~60%<30%

Lateral 2.04mm

Cortex

HippocampusCorpus callosum

Reticular thalamus

Summary• Light stimulation can activate reticular thalamus and thalamus by

activating channelrhodopsin-2 on them.• High frequency light is more effective in suppressing seizure.• Location of optic fiber is crucial, those in the reticular

thalamus(RT) and thalamus are more effective in suppressing seizure.

Acknowledgment• Dr. Shyu Bai-Chuang

• N327 members: – Chang Wei-Jen – Chang Wei-Pang

• NPAS

Thanks for your attention!!

Mechanism of seizure suppression • 130 Hz is in the range that can evoke Long-Term Potentiation

(LTP) (Bliss and Collingridge, 1993;Malenka and Nicoll, 1999). It changes synapse plasticity and network activity.

• High synaptic activity, as is induced by HFS , would then lead to downscaling of the neuronal excitability with an increase in seizure threshold as a result.

• Depolarization block occlude the kindling stimulus, which may explain why HFS has more potential than LFS.

Wyckhuys(2010). Epilepsy Research. 88, 239—246