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. 2016 Jun 22;36(25):6820–6835. doi: 10.1523/JNEUROSCI.3667-15.2016

Figure 2.

Figure 2.

Electrical stimulation of raphe enhances sensory responses in TH-positive juxtaglomerular neurons. A, Overview of GCaMP3 expression in the OB of a TH-Cre:Rosa-GCaMP3 cross. Lower image shows magnified view of glomerular layer, with individual SA somata visible. Scale bar, 50 μm. B, Odorant-evoked responses imaged from a TH-Cre:Rosa-GCaMP3 mouse. Bi, Response map showing change in fluorescence evoked by ethyl butyrate (1%, s.v.). Bii, Time course of GCaMP3 signal in two ROIs located in the caudal-lateral (1) and anteromedial (2) OB, with distinct inhalation-linked time courses similar to those seen for GAD2+ signals. C, Raphe modulation of TH+ GCaMP signal during artificial inhalation of ambient air and raphe stimulation (20 Hz, 80 pulses). Ci, Mean and SD of GCaMP signal evoked by raphe stimulation in the absence of inhalation (“no sniff”), averaged from air-responsive ROIs in five mice, showing no significant change in fluorescence during raphe stimulation and a slight increase after the end of the train attributable to hemodynamic effects. Cii, Response map showing GCaMP3 fluorescence change after raphe stimulation (same preparation as in B, with widespread increases and peak increases in distinct foci. Cii, Traces taken from two ROIs showing time course of stimulation effect. Ciii, Time course of mean and SD of GCaMP signal elicited by raphe stimulation, averaged across nine mice. As with the GAD2+ signal, raphe stimulation effects on the TH+ GCaMP signal persist for at least 8 s after stimulation ceases. D, Effect of raphe stimulus train duration on TH+ responses to air inhalation. Traces show GCaMP6f signal averaged from 10 ROIs in one mouse. Di, Responses to stimulus trains (20 Hz) of increasing duration elicit larger and longer-lasting enhancement of inhalation responses. Current intensity, 400 μA. Dii, Different preparation showing comparison of effects of an 80 pulse, 20 Hz train (0.1 ms pulse duration) and a 3 pulse, 10 Hz train (1 ms pulse duration) measured from the same ROIs. Current intensity, 250 μA. E, Traces showing raphe-stimulated increase in inhalation-linked responses in TH+ neurons during odorant inhalation. Traces show mean GCaMP6f signal averaged across 10 odorant-responsive ROIs from the same preparation, delivered during presentation of odorant (ethyl butyrate, 0.1%, s.v.), as in Figure 1. Bottom traces show an excerpt from each trial (indicated by horizontal dashed line) with the vertical scales normalized to the same inhalation response amplitude for the inhalation just before the raphe stimulus train. F, Effect of raphe stimulation on odorant-evoked RRs, measured and plotted for each of eight TH+ preparations as in Figure 1.