Enhanced BDNF signalling following chronic hypoxia potentiates catecholamine release from cultured rat adrenal chromaffin cells

Environmental stressors, including chronic hypoxia, enhance the ability of adrenomedullary chromaffin cells (AMCs) to secrete catecholamines; however, the underlying molecular mechanisms remain unclear. Here, we investigated the role of brain-derived neurotrophic factor (BDNF) signalling in rat AMCs exposed to chronic hypoxia. In rat adrenal glands, BDNF and its tropomyosin-related kinase B (TrkB) receptor are highly expressed in the cortex and medulla, respectively. Exposure of AMCs to chronic hypoxia (2% O-2; 48h) in vitro caused a significant increase to TrkB mRNA expression. A similar increase was observed in an immortalized chromaffin cell line (MAH cells); however, it was absent in MAH cells deficient in the transcription factor HIF-2. A specific TrkB agonist, 7,8-dihydroxyflavone (7,8-DHF), stimulated quantal catecholamine secretion from chronically hypoxic (CHox; 2% O-2) AMCs to a greater extent than normoxic (Nox; 21% O-2) controls. Activation of TrkB by BDNF or 7,8-DHF increased intracellular Ca2+ ([Ca2+](i)), an effect that was significantly larger in CHox cells. The 7,8-DHF-induced [Ca2+](i) rise was sensitive to the tyrosine kinase inhibitor K252a and nickel (2mm), but not the Ca2+ store-depleting agent cyclopiazonic acid. Blockade of T-type calcium channels with TTA-P2 (1m) or voltage-gated Na+ channels with TTX inhibited BDNF-induced [Ca2+](i) increases. BDNF also induced a dose-dependent enhancement of action potential firing in CHox cells. These data demonstrate that during chronic hypoxia, enhancement of BDNF-TrkB signalling increases voltage-dependent Ca2+ influx and catecholamine secretion in chromaffin cells, and that T-type Ca2+ channels play a key role in the signalling pathway.