Figure 1.

Cholinergic modulation of the glomerular circuit shapes OB output. A, Epifluorescent image showing a Lucifer yellow labeled MC. GL, Glomerular layer; MC, mitral cell; puff, puffing pipette; rec., recording electrode. B, Schematic of experimental setup showing the pipette for puffing ACh to the off-target (magenta) or on-target (black) glomerulus receiving the apical dendrite of the recorded MTC. ONL, Olfactory nerve layer. C, Current-clamp recording showing MTC responses to (arrows, 1 mm, 1 s) the “off-target” (top trace) or “on-target” (middle and bottom traces) micropuffing ACh, as illustrated in B before ACSF, and after NBQX (10 μm) and APV (50 μm). Note: off-target puffing of ACh showed no effect, while on-target puffing of ACh elicited a brief enhancement of firing followed by hyperpolarization and firing termination; and this effect was not affected by NBQX and APV. D, Typical current-clamp recording traces from one MTC showing that the nAChR blocker mecamylamine (mec.; 10 μm, red) abolished the ACh-evoked firing and reduced the following hyperpolarization in the presence of NBQX and APV (blue); the residual hyperpolarization is eliminated by the addition of scopolamine (sco.; 10 μm, green). E, Quantification data from five MTCs showing the following: (1) that the ACh-induced hyperpolarization in the presence of NBQX and APV (Control) is partially reduced by scopolamine, GBZ (10 μm), or mecamylamine, and was completely eliminated by scopolamine plus mecamylamine; and (2) that the residual hyperpolarization in the presence of scopolamine or GBZ is not affected by the combination of scopolamine and GBZ. F, Voltage-clamp recording traces from the same MTC showing that glomerular micropuffing of ACh to the target glomeruli produces an inward current in the MTC held at −60 mV in the presence of NBQX and APV (blue). The addition of mecamylamine (10 μm) in the bath blocks this inward current but reveals an outward current (red). This outward current is abolished by the further addition of scopolamine (10 μm). G, Voltage-clamp recording traces showing that glomerular micropuffed ACh-induced outward current in the presence of NBQX, APV, and mecamylamine (red) is blocked by the further addition of either scopolamine (green) or GBZ (cyan). H, Quantification data from five MTCs showing that the ACh-induced outward current shown in E is abolished by either scopolamine or GBZ. I, Typical voltage-clamp recordings showing that glomerular micropuffing of ACh (arrows) reproducibly enhances spontaneous IPSCs in MTCs (top trace). The bottom trace is a blow-up from the top trace showing the initial changes in sIPSCs in response to ACh application. J, Scatter plots showing that both the frequency (top graph) and amplitude (bottom graph) of sIPSCs in MTCs are increased by glomerular micropuffed ACh. K, Quantification data from five MTCs showing that the enhancement of both frequency (top graph) and amplitude (bottom graph) of sIPSCs by glomerular-applied ACh is eliminated by scopolamine (10 μm), but not by mecamylamine (10 μm). One-way ANOVA was performed in E, between control and sco. Ampl., Amplitude; Freq., frequency. Groups in K; repeated-measures ANOVA was performed in comparison among sco., sco. + GBZ, and sco. + GBZ + mec. groups in E. Paired t tests were performed among GBZ and GBZ + sco., mec., and mec. + sco. groups in E, as well as in H. ***p < 0.001.