Adenylate cyclase 1 promotes strengthening and experience-dependent plasticity of whisker relay synapses in the thalamus

Abstract

Synaptic refinement, a process that involves elimination and strengthening of immature synapses, is critical for the development of neural circuits and behaviour. The present study investigates the role of adenylate cyclase 1 (AC1) in developmental refinement of excitatory synapses in the thalamus at the single-cell level. In the mouse, thalamic relay synapses of the lemniscal pathway undergo extensive remodelling during the second week after birth, and AC1 is highly expressed in both pre- and postsynaptic neurons during this period. Synaptic connectivity was analysed by patch-clamp recording in acute slices obtained from mice carrying a targeted null mutation of the adenylate cyclase 1 gene (AC1-KO) and wild-type littermates. We found that deletion of AC1 had no effect on the number of relay inputs received by thalamic neurons during development. In contrast, there was a selective reduction of AMPA-receptor-mediated synaptic responses in mutant thalamic neurons, and the effect increased with age. Furthermore, experience-dependent plasticity was impaired in thalamic neurons of AC1-KO mice. Whisker deprivation during early life altered the number and properties of relay inputs received by thalamic neurons in wild-type mice, but had no effects in AC1-KO mice. Our findings underline a role for AC1 in experience-dependent plasticity of excitatory synapses.

Figure 1. Elimination of redundant whisker relay inputs proceeds normally in the ventral posteromedial nucleus of the thalamus (VPm) in the absence of adenylate cyclase 1 (AC1)

AandB, upper panels, excitatory synaptic currents recorded in a wild-type (WT;A) and a mutant VPm neuron (B) at P16 with stimulations applied to the medial lemniscus. For each neuron, EPSCs were recorded at both –70 and +40 mV with a range of stimulation intensities.AandB, lower panels show the plot of EPSC peak amplitudevs. stimulation intensity. Both neurons show all-or-none responses.C, distributions of VPm neurons receiving different numbers of relay inputs for mutant (open columns) and WT mice (grey columns) aged postnatal day (P)16–17. In both mutant and WT mice, about 80% of VPm neurons receive a single relay input.DandE, EPSCs (upper panels) and plots of EPSC amplitudevs. stimulation intensity (lower panels) from a WT (D) and a mutant VPm neuron (E) at P12.F, distribution of neurons receiving different numbers of relay inputs for mutant (open columns) and WT mice (grey columns) aged P12–13.G–Ishow the equivalent results obtained from WT and mutant VPm neurons at P7.

Figure 2. Developmental strengthening of relay synapses in the VPm was disrupted in adenylate cyclase 1 knockout (AC1-KO) mice

A–C, left panels are scatter plots of data obtained from WT (open circles) and AC1-KO neurons (grey circles) at P7, P12–13 and P16–17; the right panels show the mean values.A, developmental changes in AMPA receptor (AMPAR)/NMDA receptor (NMDAR) ratio.BandC, developmental changes in the peak amplitude of the maximal EPSCs mediated by AMPA (B; measured at –70 mV) and NMDA (C; measured at +40 mV) receptors. *P< 0.01, **P< 0.005, Mann–WhitneyUtest.

Figure 3. Reduction of quantal EPSC amplitude in AC1-KO neurons

AandB, evoked quantal EPSCs recorded in the presence of 3 mm Sr²⁺ from a WT (A) and mutant VPm neuron (B) aged P16. The scale bars apply to bothAandB.C, distributions of Sr²⁺-evoked EPSC amplitude for WT (thick grey line) and mutant neurons (thin black line). The distributions were established by pooling 100 consecutive events from each of the recorded neurons (n = 8 from 3 WT mice;n = 9 from 3 mutant mice). The inset shows the averaged Sr-EPSCs from WT (grey) and mutant neurons (black); scale bars in inset represent 4 pA and 4 ms.D, the mean peak amplitude for WT (grey column) and AC1-KO neurons (open column).P< 0.001, Mann–WhitneyUtest.

Figure 4. Synaptic release probability was not altered in mutant VPm neurons

A, paired-pulse responses from mutant (left traces) and wild-type VPm neurons (right traces) at P16.B, paired-pulse ratios of AMPA (left panel) or NMDA receptor-mediated EPSCs (right panel) at 50 and 100 ms interpulse intervals.CandD, use-dependent blockade of NMDA receptor-mediated EPSCs by MK-801 (10 μm; bath application) in a mutant (C) or heterozygous control neuron (D) at P15. Left panels show the first, third, 20th, 30th and 40th evoked EPSCs after MK-801 wash in; right panels are plots of peak amplitude of evoked EPSCs over time. Stimulation was halted during MK-801 wash in.E, similar progression of MK-801 blockade in mutant (open circles) and control neurons (filled circles). For each neuron, EPSC amplitude was normalized to the first EPSC after MK-801 wash in. Each data point represents the mean from 9 mutant neurons or 14 control neurons.

Figure 5. Sensory deprivation disrupts synaptic connectivity in thalamus of normal but not AC1-KO mice

A–Cwere obtained using WT mice.A, synaptic responses in a WT neuron recorded at P16 following whisker deprivation starting at P13.B, distributions of neurons with numbers of inputs for whisker deprived (filled columns) and untreated neurons (grey columns).C, the AMPAR/NMDAR for the whisker-deprived (filled columns) and untreated group (grey columns). **P< 0.001, Mann–WhitneyUtest.D–Fshow the equivalent results obtained using AC1-KO mice. For both the WT and the AC1-KO group, data for untreated groups (B, C, EandF) are the same as those presented in Figs 1C and 2A (P16–17, WT and AC1-KO).

Figure 6. Sensory deprivation reduces synaptic strength in the thalamus of normal but not AC1-KO mice

A–Cwere obtained from WT mice aged P14.A, EPSCs recorded at +40 and –70 mV from a neuron in deprived VPm (left panel) and another in spared VPm (right panel). Whisker deprivation was performed on one side at P13, and recordings were obtained from both sides.B, the AMPAR/NMDAR ratio of EPSCs. **P< 0.005, Mann–WhitneyUtest.C, the peak amplitude of AMPAR- and NMDAR-mediated EPSCs. *P< 0.01, Mann–WhitneyUtest.D–Fshow the equivalent data obtained from AC1-KO mice at P14.