High-Dose Melatonin and Ethanol Excipient Combined with Therapeutic Hypothermia in a Newborn Piglet Asphyxia Model

With the current practice of therapeutic hypothermia for neonatal encephalopathy, disability rates and the severity spectrum of cerebral palsy are reduced. Nevertheless, safe and effective adjunct therapies are needed to optimize outcomes. This study's objective was to assess if 18mg/kg melatonin given rapidly over 2h at 1h after hypoxia-ischemia with cooling from 1-13h was safe, achieved therapeutic levels within 3h and augmented hypothermic neuroprotection. Following hypoxia-ischemia, 20 newborn piglets were randomized to: (i) Cooling 1-13h (HT; n=6); (ii) HT+ 2.5% ethanol vehicle (HT+V; n=7); (iii) HT + Melatonin (HT+M; n=7). Intensive care was maintained for 48h; aEEG was acquired throughout, brain MRS acquired at 24 and 48h and cell death (TUNEL) evaluated at 48h. There were no differences for insult severity. Core temperature was higher in HT group for first hour after HI. Comparing HT+M to HT, aEEG scores recovered more quickly by 19h (p<0.05); comparing HT+V to HT, aEEG recovered from 31h (p<0.05). Brain phosphocreatine/inorganic phosphate and NTP/exchangeable phosphate were higher at 48hin HT+M versus HT (p=0.036, p=0.049 respectively). Including both 24h and 48h measurements, the rise in Lactate/N-acetyl aspartate was reduced in white (p=0.030) and grey matter (p=0.038) after HI. Reduced overall TUNEL positive cells were observed in HT+M (47.1 cells/mm(2)) compared to HT (123.8 cells/mm(2)) (p=0.0003) and HT+V (97.5 cells/mm(2)) compared to HT (p=0.012). Localized protection was seen in white matter for HT+M versus HT (p=0.036) and internal capsule for HT+M compared to HT (p=0.001) and HT+V versus HT (p=0.006). Therapeutic melatonin levels (15-30mg/l) were achieved at 2h and were neuroprotective following HI, but ethanol vehicle was partially protective.