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. 2018 Oct 3;4(10):eaas9426. doi: 10.1126/sciadv.aas9426

Fig. 1. Eyeblink conditioning setup and confirmation that GluR2Δ7-L7-Δγ2 mice lack both PF-PC LTD and MLI-PC FFI.

Fig. 1

(A) Mice were placed in a light- and sound-isolating chamber on a freely moving foam treadmill with their head fixed at a horizontal bar. The US consisted of a weak air puff, and the CS was a green light-emitting diode (LED) light. Eyelid movements were recorded with the magnetic distance measurement technique (MDMT). In vivo PC recordings were performed on the same treadmill system. (B) Neural circuits essential and sufficient for eyeblink conditioning, explained in detail in the main text. AIN, anterior interposed nucleus; GC, granule cells; IO, inferior olive; MLI, molecular layer interneurons; MN, motoneurons (N. III, VI, and VII); PC, Purkinje cell; PN, pontine nuclei. (C) Overview of the main known molecular mechanisms underlying PC plasticity. PF-PC LTD cascades are indicated with ocher arrows, MLI-PC cascades are indicated with brown arrows, and PF-PC long-term potentiation (LTP) and intrinsic plasticity are indicated with dashed arrows. GluR2Δ7 mice lack PF-PC LTD, L7-Δγ2 mice lack MLI-PC inhibition, and GluR2Δ7-L7-Δγ2 mice have disrupted both PF-PC LTD and MLI-PC inhibition. (D) Schematic overview of MLI-PC inhibition and PF-PC LTD induction experiments. (E) In contrast to controls, GluR2Δ7-L7-Δγ2 mice show no PF-PC LTD using an induction protocol that consisted of a 100-Hz PF stimulation in eight pulses, followed by a 110-ms delay of single CF activation at 1 Hz for 5 min. (F) Applying interstimulus intervals varying from 50 to 200 ms evoked similar levels of paired-pulse facilitation, indicating that baseline PF-PC presynaptic mediated neurotransmitter release is unaltered in GluR2Δ7-L7-Δγ2 mice. (G) GluR2Δ7-L7-Δγ2 mice show significantly less sIPSCs than controls, indicating that they indeed lack MLI-PC inhibition. For PF-PC LTD and sIPSCs, data of GluR2Δ7 and L7-Δγ2 mice are used with permission from Schonewille et al. (17) and Wulff et al. (19), respectively. AC, adenylyl cyclase; AMPAR, AMPA receptor (GluR2/3); CaMKII, Ca2+/calmodulin-activated kinase II; cAMP, cyclic adenosine monophosphate; cGMP, cyclic guanylate monophosphate; CRF, corticotropin-releasing factor; GABAA, GABA type A receptor; GABAB, GABA type B receptor; G.AP, GABAAR-associated protein; GC, guanylylcyclase; GluRδ2, glutamate receptor δ2 (GRID2); IP3, inositol trisphosphate; mGluR1, metabotropic glutamate receptor 1; NMDAR, N-methyl-d-aspartate receptor; NO, nitric oxide; PKA, cAMP-dependent protein kinase; PKC, protein kinase C; PKG, cGMP-dependent protein kinase; PLC, phospholipase C; PP1, protein phosphatase 1; PP2, protein phosphatase 2 (A+B); RAB; RAS-related protein RAB3A; SK, small conductance Ca2+-activated K+ channel; VGCC, voltage-gated Ca2+ channel. For complete statistics, we refer to table S1; *P < 0.05.