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. 2020 Nov 25;11:5974. doi: 10.1038/s41467-020-19767-w

Fig. 8. GABAergic LPBN neurons monosynaptically innervate local glutamatergic neurons.

Fig. 8

a Schematic of monosynaptic retrograde rabies virus tracing from glutamatergic LPBN neurons. b Representative images showing monosynaptic rabies spread from glutamatergic LPBN neurons in VgluT2-ires-Cre mice. Arrowheads, starter neurons infected by both TVA-eGFP (green) and rabies virus (red). The arrow indicates a GABAergic neuron (labeled with GAD1 mRNA, green) that was trans-synaptically infected by rabies virus. The third and fourth panels from the left show magnified views of the boxed areas in the second panel. Scale bars, 100 μm and 30 μm (magnified views). c Schematic of patch clamp recording from glutamatergic LPBN neurons transfected with AAV-CaMKIIα-mCherry in response to light stimulation of GABAergic LPBN neurons transfected with AAV-DIO-ChR2-eYFP. d Example image showing the expression of ChR2 in GABAergic neurons (green) and mCherry in CaMKIIα neurons (red) for targeted whole-cell patch-clamp recordings as in c. Scale bar, 30 μm. e Example of light-evoked IPSCs recorded from mCherry-positive glutamatergic neurons at a holding potential of –70 mV (left panel). The IPSCs were blocked by 1 μM TTX, and they were rescued by 100 μM 4-AP and blocked again by 10 μM bicuculline (right panel). n = 12 cells in 6 slices from 4 mice. f, g Averaged amplitude and latency of recorded IPSCs as in o. n = 15 traces recorded in 6 slices from 4 mice. h Workflow (left) and representative current clamp recording trace (right) of spontaneous spikes in an mCherry-positive LPBNCaMKIIα neuron at a holding potential of –45 mV. i Extended traces from (h) with a slow time scale showing light-induced changes in spike frequency of an LPBNCaMKIIα neuron sampled before, during, and after light stimulation of GABAergic LPBN neurons. j Time course of summarized changes in spike frequency of LPBN CaMKIIα neurons induced by light stimulation of GABAergic LPBN neurons. k Averaged firing frequency of LPBN CaMKIIα neurons during each period as indicated. All data are presented as mean ± s.e.m. and error bars represent s.e.m. *P < 0.05, **P < 0.01, and ****P < 0.0001. See also Supplementary Table 1 for further statistical information. Source data are provided as a Source Data file.