Figure 1. Optogenetic interrogation of Purkinje cell (PC) to molecular layer interneuron (MLI) circuit.

(a) Selective expression of ChR2 in cerebellar PCs. Top: Image of ChR2-YFP fluorescence (green) in a sagittal cerebellum section from PCP2-Cre; Ai32 double transgenic mice. Strong ChR2 expression was observed throughout the entire cerebellar cortex, especially in the molecular layer where PC dendrites are located. Bottom: Higher-magnification image shows expression of ChR2-YFP in PC soma (arrowhead) and dendrites. Small black holes (arrowhead) in the molecular layer represent somata of MLI that do not express ChR2-YFP. ML, molecular layer; PCL, Purkinje cell layer; GCL, granule cell layer. (b) Brief illumination (480 nm, 5 ms, 9.9 mW/mm²) evoked action potentials in ChR2-expressing PCs (top). Photostimulated PCs induced inhibitory postsynaptic currents (IPSCs) in PC-MLI cells (center) but not in stellate cells (SC; bottom). (c) Top: Photostimulation of PCs inhibited firing in a postsynaptic MLI. PC-MLI was depolarized by a depolarizing current (20 pA) to sustain action potential firing. Bottom: Activation of PCs input (at blue bar) was sufficient to briefly but completely inhibit action potential firing in postsynaptic PC-MLI cells. Points indicate means and error bars represent SEMs (n=4). (d) Superimposed traces of recordings from a connected PC and PC-MLI pair. Action potentials in a presynaptic PC (middle traces), caused by depolarizing current pulses (bottom traces), induced IPSCs in the postsynaptic PC-MLI (top traces). PC-MLI holding potential was –50 mV. (e) Distribution of IPSC latencies measured in seven PC-MLI pairs (140 trials). Mean IPSC latency was 1.2 ms with relatively low synaptic jitter (0.03 ms). (f) Distribution of amplitudes of spontaneous (blue) and evoked (red) IPSCs in PC-MLIs.