L phenyalanine

Two kinds of ACSF were used in in vitro experiments: mACSF in the dissection dish before recording and nACSF in the recording chamber. mACSF was composed of (in mM) 118 NaCl, 24 NaHCO₃, 1.5 CaCl₂, 3 KCl, 5 MgCl₂, 1.4 NaH₂PO₄, 1.3 MgSO₄, 25 d-glucose, and 1 kynurenic acid. nACSF was composed of (in mM) 122 NaCl, 25 NaHCO₃, 2.5 CaCl₂, 3 KCl, 1 MgCl₂, 0.5 NaH₂PO₄, and 12 d-glucose. Both types of ACSF were saturated with carbogen (95% O₂-5% CO₂) and maintained at pH 7.4. The drugs added to the nACSF were 5-HTP, clorgyline, pargyline, tryptamine, L-tryptophan, tyramine, L-tyrosine, 2-phenylethylamine, L-phenyalanine, NSD1015 (Sigma-Aldrich), α-methyl-5-HT, octopamine, SB206553, RX821002, GR127935, dobutamine, vasopressin (Tocris) and zolmitriptan (AstraZenica). All drugs were first dissolved as a 10 – 50 mM stock solution in water before final dilution in ACSF for in vitro ventral root reflexes recording and DIC microscopy or in saline for in vivo oxygen measurements and two photon microscopy, with the exception of zolmitriptan, which was dissolved in minimal amounts of DMSO (DMSO final concentration in nACSF was 0.04%). DMSO alone had no effect on in vitro LLR in vehicle controls, as compare to nACSF control state.