Long-term depression at olfactory nerve synapses

Abstract

The synapses formed by the olfactory nerve (ON) convey sensory information to olfactory glomeruli, the first stage of central odor processing. Morphological and behavioral studies suggest that glomerular odor processing is plastic in neonate rodents. However, long-term synaptic plasticity, a cellular correlate of functional and structural plasticity, has not yet been demonstrated in this system. Here, we report that ON-->mitral cell (MC) synapses of 5- to 8-d-old mice express long-term depression (LTD) after brief low-frequency ON stimulation. Pharmacological techniques and imaging of presynaptic calcium signals demonstrate that ON-MC LTD is expressed presynaptically and requires the activation of metabotropic glutamate receptors but does not require fast synaptic transmission. LTD at the ON--> MC synapse is potentially relevant for the establishment, maintenance, and experience-dependent refinement of odor maps in the olfactory bulb.

Figure 1.

Synaptic responses evoked by ON stimulation in olfactory glomeruli and mitral cells. A, Scheme of the recording configurations for glomerular field potentials, intracellular recordings from mitral cells, and stimulation of the olfactory nerve. GL, Glomerular layer. B, Glomerular field potentials (fEPSPs) evoked by ON stimulation. B1, The first and second rows show successive recordings in control saline (ACSF), after adding 50 μm d-APV (APV), after washout of d-APV, after adding 20 μm NBQX, after supplementing NBQX with d-APV, and after returning to NBQX (NBQX/APV wash). The third row shows the initial fEPSP component (note expanded time scale) in the presence of NBQX, after adding 1 μm TTX, and after washout of TTX. B2, Allocation of fEPSP components to presynaptic action potentials (Pre), two AMPAR-mediated postsynaptic components (AMPA1 and AMPA2), and a slow NMDAR-mediated component (NMDA). C, EPSPs induced by test stimulus to ON and recorded from the soma of MCs. First row, Successive recordings in control saline (ACSF), after adding 50 μm d-APV (APV), and after washout of d-APV. Second row, Recording in control ACSF and after adding 20 μm NBQX (note expanded time scale). The traces to the right show superposition of an AMPAR-mediated EPSP recorded in d-APV and subtraction of the control recording by the recording in NBQX.

Figure 2.

Tetanization induces long-term depression at ON synapses. A, Top, Glomerular fEPSPs during baseline recording (a) and 30 min after tetanization at 5 Hz for 20 s (b). Bottom, Time course of glomerular fEPSPs. In this and all following figures, time of tetanization is indicated by a downward arrow. Symbols represent values (mean ± SEM; n = 10) of the presynaptic component (Pre), AMPAR-mediated components (AMPA1 and AMPA2), and NMDAR-mediated component (NMDA). B, Top, ON-MC EPSPs during baseline recording (a) and 30 min after tetanization (b). Bottom, Time course of EPSPs (mean ± SEM; n = 13).

Figure 3.

ON-MC LTD saturates. Time course of glomerular fEPSPs during two tetanizations (downward arrows) at an interval of 30 min. Open triangles and filled circles represent values of presynaptic and AMPAR-mediated components, respectively (mean ± SEM; n = 5).

Figure 4.

ON-MC LTD is not dependent on NMDAR and postsynaptic calcium signaling. A, Time course of LTD of glomerular fEPSPs in control ACSF (n = 17) and in the presence of 50 μm d-APV (APV; n = 13). B, Time course of ON-MC EPSPs recorded with control patch pipette solution (n = 13) and with 20 mm BAPTA-containing pipettes (n = 10). The arrows represent the time of tetanization. Error bars represent SEM.

Figure 5.

ON-MC LTD reduces calcium transients in ON terminals. A, Calcium signals in olfactory nerve terminals loaded with a calcium indicator. Left, Fluorescence image of a dye-loaded horizontal slice, arrangement of two stimulation electrodes (S1 and S2), and an outline of two regions of interest (ROI 1 and ROI 2). ONL, Olfactory nerve layer; GL, glomerular layer; EPL, external plexiform layer. The traces show calcium signals induced in ROI 1 (blue symbols) and ROI 2 (green symbols) by stimuli delivered via S1, S2, or S1 and S2. Above each trace is a color-coded map of peak calcium signal. B, Experimental setting as above. Color-coded maps show calcium signals evoked by test stimuli delivered to S1 and S2 during baseline recordings (left) and 10-20 min after tetanization of S1 (right). C, Time course of ON Ca²⁺ signals with test stimuli delivered to S1 and S2 and tetanization delivered to S1 from the experiment illustrated in B. D, Pooled data of ON Ca²⁺ signals at tetanized (filled symbols; mean ± SEM; n = 9) and control (open symbols; n = 9) ROIs. E, Time course of glomerular fEPSPs acquired during the same set of experiments.

Figure 6.

ON-MC LTD does not require activation of AMPA or NMDA receptors and is antagonized by the mGluR antagonist MCPG. A, Time course of ON Ca²⁺ signals in control saline and in the presence of 40 μm NBQX and 50 μm d-APV (APV) with and without tetanization. B, Bath application of 1 mm MCPG (black bar; n = 14) had no effect on test fEPSPs but depressed the expression of ON-MC LTD compared with control (n = 14). C, Bath application of 1 mm MCPG (black bar; n = 14) had no effect on baseline Ca²⁺ signals but depressed the reduction of the ON Ca²⁺ signals during ON-MC LTD compared with control (n = 14). The arrows represent the time of tetanization. Error bars represent SEM.

Figure 7.

Induction of ON-MC LTD by direct activation of group I mGluRs and lack of requirement of D₂ dopamine receptors. A, Time course of ON resting Ca²⁺ concentration during bath application of the group I mGluR agonist DHPG in the continuous presence of 40 μm NBQX and 50 μm d-APV (APV). Note that DHPG reversibly decreased resting Ca²⁺ concentration (n = 11). B, Time course of ON evoked Ca²⁺ signals (top trace) and baseline Ca²⁺ concentration (bottom trace) during bath application of DHPG. Note that DHPG-induced long-lasting reduction of synaptically evoked Ca²⁺ signals and transient reduction of baseline Ca²⁺ concentration. The inset shows ON Ca²⁺ signals recorded before (1) and after (2) application of DHPG. C, Bath application of 100 μm sulpiride, a D₂ dopamine receptor antagonist (black bar; n = 5), had no effect on the expression of ON-MC LTD compared with control (n = 5). D, ON-MC LTD induced by DHPG. Time course of ON-MC EPSPs (top trace) and change in resting membrane potential (bottom trace) with bath application of DHPG. Note that DHPG-induced long-lasting reduction of ON-MC EPSPs and transient depolarization of the MCs. The inset shows an ON-MC EPSP recorded before (1) and 8 min (2) and 15 min (3) after application of DHPG. Error bars represent SEM.