INSULIN-INDUCED HYPOGLYCAEMIA INCREASES ADRENALINE SECRETION AND TYROSINE HYDROXYLASE PHOSPHORYLATION IN RAT BRAIN AND ADRENAL GLAND L Bobrovskaya, M Senthilkumaran, M Johnson University of South Australia, Sansom Institute for Health Research, Adelaide, SA 5000, Australia [email protected] Introduction Diabetes is Australia’s fastest growing chronic disease. Severe hypoglycaemia is a life- threatening complication of intensive insulin therapy for type 1 and advanced type 2 diabetes. In this study we aimed to determine the effects of insulin-induced hypoglycaemia on adrenaline secretion and tyrosine hydroxylase (TH) phosphorylation (as a marker of cell activation and catecholamine synthesis) in rat brain and adrenal gland. Methods Overnight fasted rats received a single intraperitoneal injection of insulin (10 U/kg) or saline and then were euthanised 30 min, 60 min or 90 min after injection (n=5–6 per group). Cardiac blood was collected for analysis of glucose and adrenaline. Adrenal glands and catecholaminergic brain regions were rapidly removed and TH phosphorylation at Ser40, Ser31, Ser19 and TH protein were analysed by western blotting. Results Blood glucose levels were 6–8 mmol/L in control animals versus 2 mmol/L in insulin-treated animals between 30 and 90 min. Plasma adrenaline (measured by ELISA) was significantly increased at 30 min (20 fold; p<0.05), 60 min (30 fold; p<0.001) and 90 min (13 fold; p<0.001) relative to control. pSer31TH was significantly increased in the adrenal gland at all time points (4–5 fold; p<0.01), in C1 neurons at 30 and 60 min (2 fold; p<0.05), in locus coeruleus and striatum at 60 min (2 fold; p<0.05) and in substantia nigra at 60 at 90 min (2 fold; p<0.01). pSer19TH was significantly increased in substantia nigra and striatum at 60 and 90 min (1.3–2 fold; p<0.05). Conclusion We provide evidence for the first time that TH phosphorylation in rat adrenals and brain is modulated in response to insulin-induced hypoglycaemia. The increased pSer31TH may be a mechanism to increase TH activation and catecholamine synthesis in catecholamine- producing cells in response to hypoglycaemia in vivo..