In Vivo Modulation of Sensory Input to the Olfactory Bulb by Tonic and Activity-Dependent Presynaptic Inhibition of Receptor Neurons

No effect of input magnitude or sniff frequency on the strength of GABA_B-mediated presynaptic inhibition. A, Signal-to-noise ratios (mean response amplitude/SEM; see Materials and Methods) for the glomeruli used in the analysis in B. Dotted line indicates the cutoff ratio of 3 used as a criterion for including glomeruli in the analysis. Signal-to-noise ratios were above the cutoff even for glomeruli that were at the low end of the range of response amplitudes (i.e., were weakly activated). B, Increase in odorant-evoked response amplitude after CGP35348 (1 mm) application as a function of normalized predrug response amplitude for individual glomeruli. Each filled gray circle indicates one glomerulus. Larger open squares indicate mean ± SEM after binning glomeruli into five response levels. The effect of CGP35348 is similar across bins (for statistical tests, see Results). Data are from 3 Hz sniffing experiments only and are measured after 2 s of odorant stimulation. C, Traces showing responses of one glomerulus to methyl valerate (0.75% s.v.) sampled at different artificial sniff frequencies before and after CGP35348 application. All traces are from the same glomerulus. D, Summary data showing the change in response amplitude in CGP35348 at frequencies of 1, 2, 3, and 5 Hz. Responses are measured after 2 s of odorant stimulation. Numbers in parentheses indicate number of glomeruli for each measurement. E, Traces showing the mean response to the first artificial sniff of odorant (17 glomeruli, 3 animals) before drug application (black) and in the presence of CGP35348 (red). Gray trace shows the CGP35348/predrug ratio, calculated by dividing each pair of predrug and drug traces and then averaging these ratiometric traces. The effect of CGP35348 is apparent as soon as responses are detectable and reaches peak magnitude in 50 ms or less (the ratiometric trace appears to start just before signal onset, but this corresponds to a single imaged frame and likely reflects cases in which the response becomes detectable one frame earlier in the presence of CGP35348 attributable to reduced inhibition). Response traces are normalized to the same arbitrary value; ratiometric trace is scaled for optimal visualization of time course.

No evidence for GABA_B-mediated center–surround inhibition of receptor inputs to glomeruli. A, Pseudocolor maps of calcium signals evoked by methyl valerate before (left) and after (right) application of 1 mm CGP35348, with each map normalized to its own maximum. Maps show the same data as in Figure 1A. The relative patterns are nearly identical. B, Relative distributions of response amplitudes in different glomeruli before and after CGP35348. For each experiment, responses were normalized to the maximal glomerular response, with control and CGP35348 responses normalized separately. The relative distributions are identical (for statistical tests, see Results). C, CGP35348 did not preferentially enhance responses in glomeruli with strongly activated surrounds. Plots show ratio of the effect of CGP35348 on glomerular surrounds to its effect on the glomerulus, plotted as a function of the ratio of the predrug response of a glomerulus relative to it surround. Left plot, Vehicle control; right plot, CGP35348 experiments. Lines show the fit of the function y = 1 + a log(x/b) to the data. A horizontal fit indicates no relationship between center–surround ratio and CGP35348 effects (for statistical tests, see Results).

Receptor inputs are unaffected by odorant-evoked coactivation of neighboring glomeruli. A, Pseudocolor maps of responses to BA alone (1% s.v.; left), MV alone (0.5% s.v.; middle), or the binary mixture of MV + BA (right). Maps are scaled relative to the odorant that evoked the highest response amplitude (methyl valerate). Arrow indicates BA-responsive glomerulus shown in B. MV does not evoke signals in this glomerulus but does evoke strong signals in surrounding glomeruli. B, Traces showing the calcium signal in this glomerulus in response to BA alone (red), MV alone (blue), and the binary mixture of BA + MV (black). The response to the mixture is not different from the response to BA alone. C, Summary data showing SR calculated for all glomeruli activated by only one of the odorants in the pair (dark gray bar) and for a subset of these glomeruli that were within 400 μm of a glomerulus activated by the other odorant (light gray bar). Neither of the two groups shows any significant suppression. Black bar shows responses of glomeruli activated by each odorant in the pair and shows responses to the mixture, with response amplitudes normalized to the sum of the response to each odorant alone. Responses are significantly smaller than predicted from the sum of the single-odorant responses (**p < 0.0001). ant, Anterior; lat, lateral.

Tonic presynaptic inhibition regulates receptor input to the olfactory bulb in vivo. A, Traces of presynaptic calcium signals evoked by a single artificial sniff of odorant (black) and by paired ON shocks (200 ms ISI) in a slice preparation (gray). The ON shock trace is taken from Wachowiak et al. (2005). Significant paired-pulse suppression in shock-evoked responses is apparent within the interval of the rise time of a single sniff-driven odorant response. B, ON shock-evoked responses in vivo. Images show resting fluorescence (left) and response maps (right) evoked by a single 0.1 ms (800 μA) stimulus. The stimulating electrode is visible in the resting fluorescence image. ON shock-elicited responses in glomeruli up to ∼500 μm from the stimulating electrode. Traces showing responses in two glomeruli are shown at right. C, Traces showing the effect CGP35348 (1 mm) on shock-evoked responses. Top traces show signals in glomerulus A, glomerulus B, and an average of all responsive glomeruli in this preparation, and bottom traces show responses in the same glomeruli but scaled to the same amplitude. Signals are unfiltered. ant, Anterior; lat, lateral.

GABA_B-mediated feedback inhibition of receptor inputs in vivo. A, Responses of one glomerulus to paired in vivo ON shocks delivered at ISIs from 200 to 500 ms. Traces are scaled to the same peak amplitude and bandpass filtered (low-pass Gaussian, cutoff at 18 Hz, high-pass digital τ filter, 1.4 Hz cutoff) to facilitate comparison. B, PPR (test: conditioning response amplitude) as a function of ISI before (black squares) and after (1 mm, red circles) CGP35348. PPR recovers with a time constant of ∼300 ms. CGP35348 increases the PPR at all ISIs, but significant paired-pulse suppression remains at ISIs below 600 ms. *p < 0.05 relative to predrug PPR. C, Summary of effects of CGP35348 on ON shock-evoked responses. Left, PPR before and after CGP35348 (300 ms ISI). Right, Responses to the conditioning and test ON shocks in CGP35348 relative to predrug levels (**p < 0.0001). Both conditioning and test responses increase, consistent with GABA_B-mediated tonic and feedback inhibition. D, Effects of CGP35348 on ON shock-evoked responses under isoflurane anesthesia. Whereas conditioning and test responses increase, PPR is unchanged, suggesting an absence of GABA_B-mediated feedback inhibition under isoflurane.

Feedback inhibition of receptor inputs evoked by a single sniff of odorant. A, Example of sniff–shock experiment. i, Pseudocolor maps of the response to a single artificial sniff of odorant (left) and a single ON shock (right) in the same preparation. Glomerulus 1 is activated by both the sniff and shock. Glomerulus 2 is activated only by shock, and glomerulus 3 is only activated by the odorant. ii, Traces showing the response of glomeruli 1–3 to a sniff of odorant (methyl valerate, 1% s.v.) followed ∼400 ms later by an ON shock (delivered at arrow). iii, Traces of a trial in which the shock was delivered late (∼1.8 s) after the sniff-driven odorant response. ON shocks were also delivered without odorant stimulation (data not shown). iv, Comparison of shock-evoked responses in which the shock was delivered alone (black) relative to trials in which the shock was preceded by a sniff of odorant ∼400 ms earlier (red). Traces are for glomeruli 1 and 2 from A. Glomerulus 1 shows suppressed responses after a sniff of odorant, whereas glomerulus 2 (which does not respond to the odorant) does not. B, Summary data showing the shock-evoked response after a sniff of odorant relative to the response without a preceding sniff. Data are shown for glomeruli responding to shock but not odorant (left bar) and those responding to both shock and odorant (middle, right bars). Only odorant-responsive glomeruli showed suppression after a sniff, with stronger suppression at ∼400 ms after the sniff-driven response and weak suppression at ∼1–1.8 s after the response. **p < 0.01 relative to shock-alone response. ant, Anterior; lat, lateral.