Time course and permeation of synaptic AMPA receptors in cochlear nuclear neurons correlate with input

Abstract

AMPA receptors mediate rapid glutamatergic synaptic transmission. In the mammalian cochlear nuclei, neurons receive excitatory input from either auditory nerve fibers, parallel fibers, or both fiber systems. The functional correlates of differences in the source of input were examined by recording AMPA receptor-mediated, miniature EPSCs (mEPSCs) in whole-cell voltage-clamp mode from identified neurons. Bushy, octopus, and T-stellate cells of the ventral cochlear nucleus (VCN) and tuberculoventral cells of the dorsal cochlear nucleus (DCN) receive most of their excitatory input from the auditory nerve; fusiform cells receive excitatory inputs from both the auditory nerve and parallel fibers; cartwheel cells receive excitatory input from parallel fibers alone. mEPSCs from bushy, octopus, T-stellate, and tuberculoventral cells had significantly faster decay time constants (0.35-0.40 msec) than did those from fusiform and cartwheel cells (1.32-1.79 msec). Some fusiform cells had two populations of mEPSCs with distinct time courses. mEPSCs in cells with auditory nerve input alone were inhibited by philanthotoxin, a blocker of calcium-permeable AMPA receptors, whereas mEPSCs in cells with parallel fiber input were not. Thus AMPA receptors postsynaptic to the auditory nerve differ from those postsynaptic to parallel fibers both in channel-gating kinetics and in their permeability to calcium. These results confirm the conclusion that synaptic AMPA receptors are specialized according to the source of input (Hunter et al., 1993; Rubio and Wenthold, 1997; Wang et al., 1998).

Fig. 1.

Glutamatergic inputs to cells of the cochlear nuclei. Schematic representation depicts the major sources of excitatory input to some of the cells of the cochlear nuclei. Auditory nerve fibers bifurcate to innervate cells in the unlayered ventral cochlear nucleus (VCN) and in the deep layer of the dorsal cochlear nucleus (DCN). An anterior branch of each fiber innervates bushy and T-stellate cells in the anteroventral cochlear nucleus (AVCN), and the posterior branch innervates T-stellate cells in rostral posteroventral cochlear nucleus (PVCN) and octopus cells in the most caudal and dorsal PVCN. The posterior branch continues into the deep layer of the DCN where its terminals contact dendrites of tuberculoventral and fusiform cells. Parallel fibers, the axons of granule cells that lie in clusters around the VCN and within the DCN, course dorsoventrally in the molecular layer of the DCN, innervating the dendrites of cartwheel and fusiform cells.

Fig. 2.

mEPSCs differ between cells with only auditory nerve input and those with input from parallel fibers.Left, mEPSCs appear as spontaneous inward currents of varying amplitude. Ten consecutive superimposed traces show that mEPSCs occur in all of the recorded cells. In fusiform cells some mEPSCs are rapid, whereas other mEPSCs are small and slow. In cartwheel cells mEPSCs are slow. The observation that they occur more frequently in octopus cells than in other types of cells is consistent.Right, Normalized ensemble averages of individual events from single cells (31–108 events) show that events in bushy, octopus, T-stellate, and tuberculoventral cells are more rapid than in fusiform and cartwheel cells of the DCN. The ensemble average of events in fusiform cells includes both the rapid and slow events. The time of peak was used to align all of the events to be averaged; this leads to an inflection in the rising phase of the average current in the cartwheel cell because the rise times were variable.

Fig. 3.

Amplitudes of events are variable in all cell types. The majority of mEPSCs was distinguishable from the noise (gray bars). Amplitudes varied widely within each of the cell types, on average being largest in T-stellate cells and smallest in cartwheel cells. The data are binned at 10 pA; each histogram represents data pooled from several cells of each cell type. The data are summarized numerically in Table 1.

Fig. 4.

Rise times of mEPSCs were fast in cells with auditory nerve input. The 10–90% rise times of mEPSCs from bushy, octopus, and T-stellate cells of the VCN and the tuberculoventral cell of the DCN were, on average, faster than those from fusiform and cartwheel cells of the DCN. Rise times in bushy, octopus, T-stellate, and tuberculoventral cells had narrow distributions, whereas those of fusiform and cartwheel cells had broad distributions. Rapidly rising events, events in the first 0.1 msec bin, were observed in all of the cells that receive input from the auditory nerve, but not in the cartwheel cells that do not get input from the auditory nerve. Each histogram represents data pooled from several cells of each cell type. These data are summarized numerically in Table 1.

Fig. 5.

Decay of mEPSCs in cells with auditory nerve input is faster than in cells with parallel fiber input. Single exponential decay time constants fit to individual events were generally faster in bushy, octopus, T-stellate, and tuberculoventral cells for which the excitatory input is from the auditory nerve rather than in cartwheel cells for which the excitatory input is from parallel fibers. The range of decay time constants was broad in cells with parallel fiber input, the fusiform and cartwheel cells. The data are binned at 0.2 msec. Each histogram was constructed from several cells of each cell type. The data are summarized numerically in Table 1.

Fig. 6.

Plots of rise times against amplitudes show no evidence for dendritic filtering. Plots of rise times as a function of amplitude had positive or slightly negative correlations. For each scatterplot a simple regression line was fit through the data, and the correlation coefficient was computed. Negative correlations between rise time and amplitude are indicative of filtering; an absence of correlation can be the result of the large variability in amplitude that commonly is observed in these cells. The data represent values pooled from several cells of each cell type.

Fig. 7.

Plots of rise times against decay time constants show evidence for dendritic filtering of events in some cell types. Plots of rise time against the decay time constant showed a strong positive correlation for T-stellate, tuberculoventral, fusiform, and cartwheel cells. The correlation of rise time and decay time constant are indicative of an effect on the time course of some events by dendritic filtering. For each scatterplot a linear regression line was fit through the data, and the correlation coefficient was computed. The data were pooled from several cells of each cell type.

Fig. 8.

Some fusiform cells have two populations of mEPSCs. Left, Some fusiform cells had two kinetically distinct groups of mEPSCs. Five consecutive traces are shown for each cell in which rapid events can be distinguished by eye from slow events. Right, Histograms for the decay time constant for each cell show a bimodal distribution representing two populations. The data are binned at 0.45 msec for both fusiform cells.

Fig. 9.

PhTX blocks AMPA receptors only with auditory nerve input and not with parallel fiber input. Left, The 50 μm cadmium and kainate alone had no effect on mEPSCs in a bushy cell. The amplitude, rise and decay time, and frequency of events before and after were not significantly different (p < 0.01; Student’s ttest). Center, mEPSCs were abolished completely by 50 μm PhTX in a bushy cell. Right, 50 μm PhTX did not affect mEPSCs in a cartwheel cell. The amplitude, rise and decay time, and frequency before and after were not significantly different (p < 0.01; Student’s t test). Strychnine (1 μm) was present in the bath for the duration of the recording to isolate the mEPSCs. All traces were filtered off-line at 5 kHz for presentation purposes.

Fig. 10.

Cells with auditory nerve input alone have rapidly gating calcium-permeable AMPA receptors. Plotting the average decay time constant (±SEM) versus the average amplitude of events after PhTX relative to before shows that those cells with rapidly decaying mEPSCs (bushy, T-stellate, octopus, and tuberculoventral cells) are also sensitive to PhTX. Those cells with slower mEPSC decays (fusiform and cartwheel cells) were insensitive to PhTX. Thex-axis was calculated from average amplitude values for each cell. The decay time constants represent the average decay of events in each cell before the application of PhTX, kainate, and Cd²⁺. The amplitudes were not changed significantly in fusiform and cartwheel cells (p < 0.01; Student’s t test) but were not identical before and after PhTX (Amplitude_PhTX/Amplitude_Normal ≠ 1).