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Neurotransmitters, Gliotransmitters, and Modulators

開催日 2014/9/11
時間 16:00 - 17:00
会場 Poster / Exhibition(Event Hall B)

Comparison of the characteristics of two different fluorescent probes for neurotransmitter release

  • P1-016
  • 引間 卓弥 / Takuya Hikima:1 アーバスノット ゴードン / Gordon W Arbuthnott:1 
  • 1:沖縄科学技術大学院大学 行動の脳機構ユニット / Brain Mechanisms for Behaviour Unit, Okinawa Institute of Science and Technology, Okinawa, Japan 

Cortical synapses are important for fundamental brain function and changes in synaptic strength affect how brain processes information. Typically an increase in synaptic efficacy is accompanied by an increase in presynaptic neurotransmitter release and imaging methods are an effective tool to investigate the kinetics for neurotransmission in cortical presynaptic terminals. We used two fluorescent probes for neurotransmitter release, synaptophysin-pHluorinx2 (SypHx2) and intensity-based glutamate-sensing fluorescent reporter (iGluSnFR) to optically visualize neurotransmitter release. SypHx2 is the fusion of synaptic vesicle protein synaptophysin with two molecules of the super ecliptic pHluorin, pH-sensitive green fluorescent protein (GFP), iGluSnFR is a sensor for synaptically released glutamate constructed using a bacterial periplasmic binding protein and circularly permulated GFP. We could detect the fluorescent change of SypHx2 (ΔSypHx2) with 5 pulses electrical stimulation and estimate the size of the readily releasable pool (RRP) after high frequency stimulation. On the other hand, the fluorescent change of iGluSnFR (ΔiGluSnFR) was much higher. ΔiGluSnFR signals to single pulse field stimulation were reliably detected and reached a plateau after 5 pulses. Both signals were dependent on external calcium concentration. We found that cAMP/PKA activity uncovered an increased number of presynaptically silent synapses using both methods, and the increase depended on the size of the RRP. Based on our results, these two fluorescent probes are useful tools to follow the alteration of the presynaptic efficacy and examine the properties of presynaptic sites more precisely.

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