Apetx2

Agmatine sulfate salt (Cat#a7127), histamine (Cat#y0001779), compound 48/80 (Cat#C2313), resiniferatoxin (RTX, Cat#R8756), chlorpheniramine maleate (Cat#C3025), chloroquine (CQ, Cat#C6628), naloxone hydrochloride (Cat#N3136), amiloride hydrochloride (Cat#1019701) were all obtained from Sigma-Aldrich (St. Louis, MO, United States). MC903 (Cat# 2700/10) was purchased from R&D systems (Minneapolis, MN, United States). Morphine hydrochloride was obtained from China Northeast Pharmaceutical Group Shenyang No. 1 Pharmaceutical Co., Ltd (Shenyang City, China). naloxone hydrochloride HC-030031 (Cat#2896), capsazepine (CPZ, Cat#0464) and APETx2 (Cat#4804) were purchased from Tocris (Minneapolis, MN, United States). ZnCl₂ was obtained from National Pharmaceutical Group pharmaceutical Co., Ltd (Beijing, China). RTX and HC-030031 were freshly dissolved in 10% DMSO. Other reagents were dissolved in saline. Information relating to timing and doses are indicated in the results section or figure legends.

Spinal cord-notochord preparations (10–20 segments; n=28) were dissected from the area between the gills and the dorsal fin as described previously53 (link) and pinned down in a Sylgard-lined chamber continuously perfused with oxygenated physiological solution which was kept at 8–10 °C. NMDA (100–150 μM, Tocris) was added to the physiological solution to induce fictive locomotion30 (link). The motor activity was monitored by the use of glass suction electrodes positioned on two opposite ventral roots (see Fig. 6b). Swimming sequences were analysed for at least 20 cycles recorded during each experimental condition. The effects on the locomotor burst frequency induced by decreases in extracellular pH (pH 6.9 and 6.5) were analysed, as well as the effects of somatostatin (10 nM to 1 μΜ, Tocris). Corresponding experiments were also performed in the presence of the ASIC3 antagonist APETx2 (1 μM) or the somatostatin receptor sst₂ antagonist CYN-154806 (2 μM; Tocris).