Involvement of A5/a7 Noradrenergic Neurons and B2 Serotonergic Neurons in Nociceptive Processing: A Fiber Photometry Study

Abstract In nociceptive processing, the descending antinociceptive system (DAS) is well known to include a number of descending pathways.As the main pathway of DAS, the locus coeruleus (A6) – spinal horn circuit including the noradrenergic (NAergic) system andthe periaqueductal gray-rostral ventromedial medulla (B3) – spinal horn circuit including the serotonergic (5-HTergic) system are well known.OtherNAergic or 5-HTegic systems, however,are less well known. Some studies have reported that A5/A7 noradrenaline (NA) neurons or B2 serotonin (5-HT, 5-hydroxytryptamine) neurons partially participate in DAS.In this study, we recorded G-CaMP6 green fluorescence signal intensities in theA5/A7/B2 sites of awake mice in response to acute tail pinch stimuli, acute heat stimuli, and in the context of a noninvasive control test, using fiber photometry with acalcium imaging system.We first introduced G-CaMP6 in the A5/A7 NA neuronal soma or B2 5-HT neuronal soma, using transgenic mice carrying tetracycline-controlled transactivator transgene (tTA) under the control of either a dopamine β-hydroxylase promoter or a tryptophan hydroxylase-2 and by the site-specific injection of adeno-associated virus (AAV-TetO(3G)-G-CaMP6).The specific expression patterns of G-CaMP6 were clearly confirmed in the A5/A7 areasand B2 area.After confirming the specific expression patterns of G-CaMP6, we recorded G-CaMP6 green fluorescence signals in these sites fromawake mice in response to acute nociceptive stimuli.G-CaMP6 fluorescence intensity in the A5, A7, and B2 groups wasrapidly increased by acute nociceptive stimuli,soon returning to baseline fluorescence intensity.This was not observed in the context of the noninvasive control test.Weused mCherry red fluorescent protein as a control indicator. mCherry fluorescence intensity was not significantly changed by acute nociceptive stimuli or in the noninvasive control test in any of the groups. These results clearly indicate that acute nociceptive stimuli rapidly increase the activities of A5/A7 NA neurons or B2 5-HT neurons, suggesting that A5/A7 NA neurons or B2 5-HT neurons play important roles in nociceptive processing in the central nervous system.