In these experiments, we used full-field, high-intensity light to stimulate a maximal number of MLIs while recording simultaneously from both a Golgi cell and a nearby Purkinje cell (Figure 7A). Light pulses evoked large inhibitory synaptic currents in all recorded PCs, which is consistent with the activation of many

MLIs (Figures 7C and 7D; see Experimental Procedures). These synaptic responses were eliminated by the GABAA-receptor antagonist gabazine. In contrast, even though many MLIs were activated in these experiments, we never observed any synaptic input onto simultaneously recorded Golgi cells (n = 6). Previous studies have also suggested selleck kinase inhibitor that MLIs and Golgi cells are gap junction coupled (Sotelo and Llinás, 1972). We therefore tested for such connections but found no electrical coupling between any MLIs and Golgi cells in 31 paired recordings (mean junctional conductance = −0.01 ± 0.01 nS). These experiments, along with the lack of synaptic connections observed in paired recordings and with ChR2 stimulation, suggest that despite the many MLIs in the molecular layer in close proximity to Golgi

cell dendrites, MLIs do not make fast inhibitory synapses or gap junctional connections onto Golgi cells. These findings change the inhibitory wiring diagram of the cerebellar cortex by establishing that Golgi cells are inhibited by other Golgi cells and not by MLIs (Figure 8A), but what see more are the consequences of this circuit revision? MF activation evokes IPSCs that arrive earlier onto Golgi cells than onto Purkinje cells (Figure 2). To determine the implications for Golgi cell activity, we examined the timing of inhibition relative to excitation in these cells. MF activation should excite Golgi cells directly (MF→Golgi cell) as well as indirectly by activating granule cell synapses (MF→granule cell→Golgi cell). Indeed, we find that brief, high-intensity optical stimulation of MFs can evoke EPSCs onto SB-3CT Golgi cells that consist of

two discrete components (Figure 8B). Through the use the CB1 receptor agonist WIN 55,212-2 (WIN), which is known to suppress release from granule cells onto Golgi cells (Beierlein et al., 2007), we found a selective reduction of the second component of the EPSC following ChR2 activation (EPSC1: 2% ± 4% reduction, p = 0.79; EPSC2: 43% ± 6% reduction, p < 0.001, n = 7; Figures 8B and 8C). The observed delay between EPSC1 and EPSC2 and the pharmacological sensitivity of EPSC2 establishes that the second component of the EPSC is a result of disynaptically activating granule cell synapses. We then compared the relative timing of evoked IPSCs and EPSCs. These experiments revealed that disynaptic inhibition from Golgi cells and disynaptic excitation from granule cells arrive simultaneously (Δt = 0.1 ± 0.3 ms, n = 11, p = 0.8; Figure 8D). This is very different from the timing of excitation and inhibition for Purkinje cells (Figure 8E).