Fig. 1

Nociceptive stimuli increased G-CaMP6 fluorescence intensity of the A13 dopaminergic cells in the zona incerta. a A genetic scheme of neuron-specific expression of fluorescent proteins. G-CaMP6 and mCherry fluorescent proteins were expressed by a DAT promotor-driven Cre recombinase. b AAV injection into the zona incerta site. c The confirmation of proper fiber positioning. The silica fiber was just above the zona incerta site. Scale bar: 300 μm. d G-CaMP6 and mCherry were specifically expressed in A13 dopaminergic neuronal soma. G-CaMP6/mCherry-positive neurons were rarely observed outside the A13 area. Tyrosine hydroxylase (TH)-positive cells were found in the A13 area and 92.0% of them expressed G-CaMP6 (n = 3). All G-CaMP6-positive cells also expressed mCherry, and 92.2% of them expressed TH. Scale bar: 100 μm. e and f Averaged traces of fluorescence intensity of G-CaMP6 and mCherry. Each trace is the average of six mice. e G-CaMP6/mCherry (green/red) traces in the pinch group and G-CaMP6/mCherry (blue/purple) in the touch group. f G-CaMP6/mCherry (green/red) traces in the heat group and G-CaMP6/mCherry (blue/purple) in the low heat group. The horizontal bar represents the time of stimulation. Vertical bars indicate S.E.M. g Effects of acute nociceptive stimuli and non-nociceptive stimuli on G-CaMP6 fluorescence intensity. h and i Characteristics of G-CaMP6 fluorescence in response to acute nociceptive stimuli. Onset latency: time from the start of stimulus to the time when fluorescence signal intensity exceeded the maximum value during the baseline period. Peak latency: time from the start of stimulus to the time when fluorescence signal intensity arrived at the maximum point. All data in g-i are expressed as values of the mean ± S.E.M and were analyzed with two-way factorial ANOVA with Sidak’s post-hoc test (n = 6, each). P values show the results of Sidak’s test