Augmentation of glycolytic metabolism by meclizine is indispensable for protection of dorsal root ganglion neurons from hypoxia-induced mitochondrial compromise

Abstract

To meet energy demands, dorsal root ganglion (DRG) neurons harbor high mitochondrial content, which renders them acutely vulnerable to disruptions of energy homeostasis. While neurons typically rely on mitochondrial energy production and have not been associated with metabolic plasticity, new studies reveal that meclizine, a drug, recently linked to modulations of energy metabolism, protects neurons from insults that disrupt energy homeostasis. We show that meclizine rapidly enhances glycolysis in DRG neurons and that glycolytic metabolism is indispensable for meclizine-exerted protection of DRG neurons from hypoxic stress. We report that supplementation of meclizine during hypoxic exposure prevents ATP depletion, preserves NADPH and glutathione stores, curbs reactive oxygen species (ROS) and attenuates mitochondrial clustering in DRG neurites. Using extracellular flux analyzer, we show that in cultured DRG neurons meclizine mitigates hypoxia-induced loss of mitochondrial respiratory capacity. Respiratory capacity is a measure of mitochondrial fitness and cell ability to meet fluctuating energy demands and therefore, a key determinant of cellular fate. While meclizine is an ‘old’ drug with long record of clinical use, its ability to modulate energy metabolism has been uncovered only recently. Our findings documenting neuroprotection by meclizine in a setting of hypoxic stress reveal previously unappreciated metabolic plasticity of DRG neurons as well as potential for pharmacological harnessing of the newly discovered metabolic plasticity for protection of peripheral nervous system under mitochondria compromising conditions.

Hypoxia potentiates the capacity of melanoma cells to evade cisplatin and doxorubicin cytotoxicity via glycolytic shift

“…6). Cultures challenged with drugs under normoxic and hypoxic conditions were processed in parallel and XF24‐implemented sequential additions of mitochondrial effectors, oligomycin, FCCP, 2‐deoxyglucose (2DG), and antimycin A, served to determine the treatments signatures on respiratory parameters, as we previously described [16,26]. Under normoxic conditions, baseline OCR in YUMM1.7 was ~ 30% lower compared to B16F10 cells.…”

“…The B16F10 cell line (ATCC CRL‐6475) [15] was cultured in DMEM (Invitrogen # 11965092) with 10% FBS, and NIH3T3 embryonic fibroblast cell line (ATCC CRL‐1658) was cultured in DMEM with 10% bovine calf serum (ATCC #30‐2030). Hypoxia treatments were done as described [16]; briefly, cultures were placed in a gas‐tight modular incubator chamber (Billups Rothenberg, Del Mar, CA) and flushed for 3 min with 5% CO 2 balanced with 95% nitrogen at a flow rate of 50 L·min −1 . The oxygen content in the chamber was 0.8 ± 0.2% measured by the oxygen pen 800047 (Sper Scientific, Scottsdale, AZ, USA) with water container placed in the chamber to maintain a humidified environment.…”

“…B16F10 and YUMM1.7 cells were seeded on coverslips in 24‐well plates, treated as indicated and fixed in 4% paraformaldehyde [16,21]. Cells were permeabilized with 0.1% Triton X‐100/0.1% sodium citrate in PBS and blocked with 3% BSA (w/v)/1% donkey serum (v/v) in PBS followed by mouse anti‐Cox1 (1: 1500) antibody (Invitrogen‐459600).…”

“…Oxygen consumption rates (OCR) were measured using XF24 extracellular flux analyzer (Seahorse, Agilent, Folsom, CA, USA) according to established protocols [23–25] and as we described [16,21,26]. B16F10 and YUMM1.7 cells were seeded in XF24 plates ([0.8‐1.2] × 10 4 /well), grown overnight, and treated as indicated.…”

A New Method for Primary Culture of Mouse Dorsal Root Ganglion Neurons

“…Spinal DRGn are afferent neurons of peripheral nervous system and primary neurons of sensory conduction system. With the increasing research on the nervous system, such as the pain mechanism of diseases [1][2][3], spinal cord injury repair [4,5], electrophysiological research [6][7][8][9], it is necessary to establish a simple and easy-to-obtain primary nerve cell culture scheme.…”