Figure 7. Effects of changes in [Ca²⁺]_o and ISP on depression of GABA release, the size of the RRP and refilling of neurotransmitter pools at BC–PC synapses following tetanic stimulation.

A, sample records of eIPSCs evoked by tetanic stimulation (20 Hz for 1.5 s) at 1.5 (upper) and 5 mm[Ca²⁺]_o (lower) before (black) and during application of 20 μm ISP (red). Each record was obtained by averaging five successive traces. B, the amplitude of eIPSCs evoked by repetitive stimulation as in A at different levels of [Ca²⁺]_o and expressed as relative amplitude to the initial IPSCs in each [Ca²⁺]_o before (○) and during the application of ISP (•) (n = 5–6). C, estimation of the RRP from the relationship between the cumulative eIPSC amplitude (determined in B) and the pulse number during tetanic stimulation. Cumulative curves were compiled from the amplitudes of successive IPSCs during repetitive stimulation normalized to the first IPSC amplitude recorded in each trial in 5 mm[Ca²⁺]_o. Data points during a steady-state phase (from the 21st to the 30th and the 16th to the 20th pulses for 20 Hz, and 50 Hz stimulation, respectively) were fitted by linear regression, and extrapolated to the y-axis, giving values of the RRP size in the control solution (○ for 20 Hz, ▵ for 50 Hz, and lines in black). ISP (20 μm) significantly increased the RRP size (• and line in red). D, pooled data of the PPR sizes in the absence (open column) and the presence of ISP (grey column) in 2.5 and 5 mm[Ca²⁺]_o, and the pool size determined with 50-Hz tetanic stimulation. ISP (20 μm) significantly increased the size of the RRP under both [Ca²⁺]_o conditions (2.5 mm[Ca²⁺]_o, P < 0.05, n = 8; 5 mm[Ca²⁺]_o, P < 0.01, n = 8). E, effects of ISP on the rate of transmitter refilling. After conditioning with high frequency stimulation (20 Hz, 30 pulses) in 5 mm[Ca²⁺]_o, single test stimuli with different intervals were applied to evoke IPSCs in a PC and superimposed before (upper trace) and during ISP (20 μm) application (lower trace). ISP accelerated the refilling process. F, pooled data of averaged time course for the recovery of synaptic depression after the tetanic stimulation (20 Hz, 30 pulses) in the control solution (○, n = 10) and in the presence of 20 μm ISP (•, n = 9). Smooth curves represent single exponential fits to the averaged data. ISP application significantly shortened the time constant of IPSC recovery from 210.4 ± 21.4 ms (n = 10) to 86 ± 11.6 ms (n = 9) (P < 0.05, unpaired t test).