Activation of postsynaptic GABAB receptors modulates the bursting pattern and synaptic activity of olfactory bulb juxtaglomerular neurons

Abstract

Olfactory bulb glomeruli are formed by a network of three major types of neurons collectively called juxtaglomerular (JG) cells, which include external tufted (ET), periglomerular (PG), and short axon (SA) cells. There is solid evidence that gamma-aminobutyric acid (GABA) released from PG neurons presynaptically inhibits glutamate release from olfactory nerve terminals via activation of GABA(B) receptors (GABA(B)-Rs). However, it is still unclear whether ET cells have GABA(B)-Rs. We have investigated whether ET cells have functional postsynaptic GABA(B)-Rs using extracellular and whole cell recordings in olfactory bulb slices. In the presence of fast synaptic blockers (CNQX, APV, and gabazine), the GABA(B)-R agonist baclofen either completely inhibited the bursting or reduced the bursting frequency and increased the burst duration and the number of spikes/burst in ET cells. In the presence of fast synaptic blockers and tetrodotoxin, baclofen induced an outward current in ET cells, suggesting a direct postsynaptic effect. Baclofen reduced the frequency and amplitude of spontaneous EPSCs in PG and SA cells. In the presence of sodium and potassium channel blockers, baclofen reduced the frequency of miniature EPSCs, which were inhibited by the calcium channel blocker cadmium. All baclofen effects were reversed by application of the GABA(B)-R antagonist CGP55845. We suggest that activation of GABA(B)-Rs directly inhibits ET cell bursting and decreases excitatory dendrodendritic transmission from ET to PG and SA cells. Thus the postsynaptic GABA(B)-Rs on ET cells may play an important role in shaping the activation pattern of the glomeruli during olfactory coding.

FIG. 1

Effects of baclofen on spontaneous spike bursting in external tufted (ET) cells. Extracellular recordings were made from an ET cell exhibiting bursts of actions potentials that persisted in the presence of blockers of fast synaptic transmission [6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), 10 μM; (±)-2-amino-5- phosphopentanoic acid (APV), 50 μM; gabazine, 10 μM]. The burst parameters are presented by different colors in the graphs with corresponding scales. Baclofen reduced the frequency of bursts and the firing frequency but it increased the burst duration and the number of spikes/burst. These effects were reversed after addition of the μ -aminobutyric type B receptor (GABA_B-R) blocker CGP55845 to the bath (shown by the double bars above). The traces were smoothed by adjacent averaging from 30 consecutive data points. The timescale is common for all graphs. Bottom: 4 representative traces of extracellular recordings in each condition.

FIG. 2

Modulation of intraburst properties by baclofen. A, top: scatter plot of the interspike intervals (ISIs) calculated from the beginning of first spike in the burst under different pharmacological conditions. Bottom: sliding color-coded autocorrelograms (triggered on the first spike in the burst) performed every minute using 2-min sampling periods. Application of 10 μM CNQX, 50 μM APV, and 10 μM gabazine increased the regularity of ISIs within the burst as shown by the decrease in the variability of the data points that show higher density around regularly spaced peaks. Additional application of baclofen reduced the ISIs and introduced additional peaks due to the recruitment of more spikes. This effect was reversed by the GABA_B-R blocker CGP55845. B: autocorrelograms of ISIs within bursts constructed using spike occurrence in 2-min sampling periods under different pharmacological conditions. The autocorrelograms were normalized to the bin width, the sampling period, the firing frequency, and the interburst frequency (see METHODS). C: evaluation of the slopes of the ISIs as a function of the interval number within a burst. The slopes were calculated using linear regression fit under different pharmacological conditions. All data obtained in this figure were obtained from the same cell.

FIG. 3

Direct inhibitory postsynaptic effects of baclofen on ET cells. A: current-clamp recording from an ET cell showing the inhibitory effect of baclofen in the presence of the blockers of fast synaptic transmission (CNQX, 10 μM; APV, 50 μM; and gabazine, 10 μM). Bottom 4 traces: bursting activity at extended timescale under different pharmacological conditions as indicated in the first trace. The fast synaptic blockers eliminated the synaptic potentials that were mostly obvious between bursts in control condition. Baclofen slightly hyperpolarized the membrane potential and significantly decreased and eliminated the bursting frequency. This effect was reversed by addition of the GABA_B-R blocker CGP55845. B: another ET cell was recorded in voltage clamp at holding potential (HP) = −60 mV in the presence of tetrodotoxin (TTX, 1 μM) and blockers of fast synaptic transmission (CNQX, 10 μM; APV, 50 μM; gabazine, 10 μM). Baclofen induced an outward current that was reversed by the GABA_B-R receptor antagonist CGP55845. C: plot of the current–voltage relationship in the same ET cell as in B under different pharmacological conditions. Baclofen produced an outward current at membrane potentials above −90 mV and an inward current at membrane potentials below −90 mV, which is near the reversal potential of potassium channels. Inset: current response to a 50-mV hyperpolarizing voltage step from −60 to −110 mV (duration 0.5 s) under different pharmacological conditions. During baclofen application, a larger current (dotted trace) was produced in response to the same voltage step, indicating an increase in conductance (i.e., decrease in resistance) of the cell in response to GABA_B-R activation.

FIG. 4

Baclofen effects on spontaneous and evoked bursts of excitatory postsynaptic currents (EPSCs) in periglomerular (PG) cells. All traces were obtained from the same PG cell recorded in voltage clamp at HP = −60 mV. Baclofen blocked almost completely the spontaneous bursts of EPSCs and the olfactory nerve–evoked bursts of EPSCs were either eliminated (failure, not shown) or reduced in amplitude. Baclofen also increased the latency of the EPSC bursts. These effects were reversed after addition of the GABA_B-R blocker CGP55845 to the bath. The spontaneous and evoked EPSCs were blocked by the non–N-methyl-D-aspartate receptor blocker CNQX. A partial recovery of the EPSCs was obtained after washout of the drugs.

FIG. 5

Baclofen reduces the frequency of action potential–independent EPSCs [miniature (m)EPSCs]. A, top: current trace of a short axon (SA) cell recorded in voltage clamp at HP = −70 mV with inward deflections, indicating the occurrence of spontaneous EPSCs. The pipette intracellular solution contains cesium and QX-314 (see methods for details). Bottom: corresponding histogram of the frequency of EPSCs throughout the experiment. The histogram was first constructed using a bin width of 10 s (thin line), and then it was further smoothed using adjacent averaging of 5 data points. The histogram was normalized to the bin width to reflect the mean frequency of EPSCs. Note that the increase or decrease in EPSCs was associated with a slight inward or outward current, respectively, suggesting that background EPSC frequency level might contribute to the mean current level. B: sample current traces shown at extended timescale (3 s each) under different pharmacological conditions corresponding to the recording shown in A. Application of the blockers (TTX, 1 μM; gabazine, 10 μM; TEA, 10 mM; barium chloride, 1 mM) significantly increased the EPSC frequency, which became highly irregular. The TTX-resistant mEPSCs were inhibited by baclofen and this effect was reversed by additional application of the GABA_B-R blocker CGP55845. Further addition of the calcium channel blocker cadmium blocked most of the mEPSCs.